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Phylogeny and taxonomy of root-inhabiting Cryptosporiopsis species, and C. rhizophila sp. nov., a fungus inhabiting roots of several Ericaceae.

Three Cryptosporiopsis species have thus far been isolated from roots of woody plants. A fourth species, which was recently isolated from roots of Calluna vulgaris, Erica tetralix, Vaccinium vitis-idaea, and V. myrtillus in The Netherlands, is described here as new. Sporulation on the natural substratum has not been observed and the morphological description of this fungus is therefore based on characters expressed on oatmeal and malt extract agars. The phenotypic characters indicated a close relationship with the other root-inhabiting species of Cryptosporiopsis and species of the associated teleomorph genus Pezicula. This relationship was confirmed by phylogenetic analyses using sequence data of the 5.8S nuclear rDNA and flanking internal transcribed spacers. In order to facilitate recognition of this possibly under-recognized category of root inhabitants, a key to the root-inhabiting Cryptosporiopsis species based on characters in vitro is given.

Ascomycota↗

Using unnatural protein fusions to engineer resveratrol biosynthesis in yeast and Mammalian cells.

Resveratrol is a naturally occurring defense compound produced by a limited number of plants in response to stresses. Besides cardiovascular benefits, this health-promoting compound has been reported to extend life spans in yeasts, flies, worms, and fish. To biosynthesize resveratrol de novo, tyrosine ammonia lyase (TAL), 4-coumarate CoA-ligase (4CL), and stilbene synthase (STS) were isolated from Rhodobacter sphaeroides, Arabidopsis thaliana, and Vitis vinifera, respectively. Yeast cells expressing 4CL and STS produce resveratrol when fed with 4-coumaric acid, the substrate of 4CL. When a translational fusion protein joining 4CL and STS was used, yeast cells produced 15-fold more resveratrol than the cotransformed cells, suggesting that physical localization of 4CL and STS facilitate resveratrol production. When the resveratrol pathway was introduced into human HEK293 cells, de novo biosynthesis was detected, leading to intracellular accumulation of resveratrol. We successfully engineered an entire plant natural product pathway into a mammalian host.

Acyltransferases↗

Analysis of proanthocyanidin cleavage products following acid-catalysis in the presence of excess phloroglucinol.

The analysis of proanthocyanidin cleavage products after acid-catalysis in the presence of excess phloroglucinol was investigated. In the developed analytical method, a solution of 0.1 N HCl in methanol, containing 50 g/L phloroglucinol and 10 g/L ascorbic acid was prepared. The proanthocyanidin of interest was reacted in this solution (5 g/L) at 50 degrees C for 20 min, and afterward combined with 5 volumes of 40 mM aqueous sodium acetate before analysis by reversed-phase HPLC using an aqueous acetic acid and methanol gradient. This procedure was used to investigate the composition of proanthocyanidins isolated from the seed and skin tissue of Vitis vinifera L. berries. The results compared favorably to results obtained when benzyl mercaptan was used as the trapping nucleophile, indicating that phloroglucinol is an effective reagent for this analysis.

Anthocyanins↗

Changes in nitrogen compounds in must and wine during fermentation and biological aging by flor yeasts.

Urea, ammonium, and free amino acid contents were quantified in a must from Vitis vinifera cv. Pedro Ximenez grapes and in fermented wine and after a short aging of this wine by Saccharomyces cerevisiae race capensis yeast under variable oxygen availability conditions. The previous compounds were also determined in a wine in which the nitrogen source was depleted by the same race of flor yeast (old wine) and also following the addition of ammonium ion, L-glutamic acid, and L-proline. Under specific conditions such as low oxygen level and the absence of some nutrients, the yeasts release some amino acids including L-threonine, L-tryptophan, L-cysteine, and L-methionine to the medium. These amino acids must originate primarily in a de novo synthesis from ethanol that regenerates NAD(P)+. On the basis of these results, the yeasts may be able to use amino acids not only as nitrogen sources but also as redox agents to balance the oxidation-reduction potential under conditions of restricted oxygen, when electron transport along the respiratory chain may be hindered or limited.

Aging↗

Determination of anthocyanidins in berries and red wine by high-performance liquid chromatography.

A high-performance liquid chromatographic (HPLC) method for the determination of anthocyanidins from berries and red wine is described. Delphinidin, cyanidin, petunidin, pelargonidin, peonidin, and malvidin contents of bilberry (Vaccinium myrtillus), black currant (Ribes nigrum), strawberry (Fragaria ananassa cv. Jonsok), and a Cabernet sauvignon (Vitis vinifera) red wine were determined. The aglycon forms of the anthocyanins present in the samples were revealed by acid hydrolysis. A reversed phase analytical column was employed to separate the anthocyanidins before identification by diode array detection. The suitability of the method was tested by determining the recovery (95-102% as aglycons and 69-104% from glycosides) for each anthocyanidin. Method repeatability was tested by charting the total aglycon content of two samples over a period of 14 analyses and determining the coefficients of variation (1.41% for bilberry and 2.56% for in-house reference material). The method developed proved thus to be effective for reliable determination of anthocyanidins from freeze-dried berry samples and red wine. The total anthocyanidin content of the tested samples was as follows: in-house reference material, 447 +/- 8 mg/100 g; strawberry, 23.8 +/- 0.4 mg/100 g; black currant, 135 +/- 3 mg/100 g; bilberry, 360 +/- 3 mg/100 g; and Cabernet sauvignon red wine, 26.1 +/- 0.1 mg/100 mL.

Anthocyanins↗

Antioxidant properties of trans-epsilon-viniferin as compared to stilbene derivatives in aqueous and nonaqueous media.

trans-Epsilon-viniferin, the dimer of resveratrol, extracted from Vitis vinifera, has been evaluated for its antioxidant capacity. Its properties have been compared to those of resveratrol and synthetic stilbenic derivatives (4-hydroxystilbene, 4,4'-dihydroxystilbene, 3,5-dihydroxystilbene, and trimethylresveratrol), in regard to their liposolubility using two media with different polarity. The bleaching of beta-carotene by lipoperoxyl (LOO.) radicals in an oil/water (O/W) emulsion and the scavenging of superoxide anions (O(-)(2) in dimethyl sulfoxide (DMSO) using 5,5-dimethyl-1-pyrroline-N-oxide as a spin trap were followed using UV-visible and electron paramagnetic resonance, respectively. Epsilon-viniferin exhibits the best antioxidant capacity in the DMSO/O(-)(2) polar system (IC(50) = 0.14 mM) while 4,4'-dihydroxystilbene presents the highest antioxidant capacity in the O/W/LOO. system (inhibition of beta-carotene bleaching, 82%). Partition coefficients and kinetics of partition between 1-octanol and water were measured to discuss the antioxidant efficiency of the compounds in relation with their chemical structure.

Antioxidants↗

Composition of grape skin proanthocyanidins at different stages of berry development.

The composition of grape (Vitis vinifera L. cv. Shiraz) skin proanthocyanidins has been determined at different stages of berry development. Beginning approximately 3 weeks after fruit set and concluding at commercial ripeness, the composition of isolated skin proanthocyanidins was determined using the following analytical techniques: elemental analysis, UV-Vis absorption spectroscopy, reversed-phase HPLC after acid-catalysis in the presence of excess phloroglucinol, gel permeation chromatography, electrospray ionization mass spectrometry (ESI-MS), and (13)C NMR. On the basis of these analyses, berry development was correlated with an increase in proanthocyanidin mean degree of polymerization, an increase in the proportion of (-)-epigallocatechin extension subunits, and increases in the level of anthocyanins associated with the proanthocyanidin fraction. Additionally, data acquired from ESI-MS of the isolates following acid-catalysis in the presence of excess phloroglucinol is consistent with pectin-bound proanthocyanidins.

Anthocyanins↗

"Untypical aging off-flavor" in wine: synthesis of potential degradation compounds of indole-3-acetic acid and kynurenine and their evaluation as precursors of 2-aminoacetophenone.

Kynurenine (1) and indole-3-acetic acid (2) are considered as potential precursors of 2-aminoacetophenone (3), which is regarded to be the aroma impact compound causing an "untypical aging off-flavor" (UTA) in Vitis vinifera wines. The mechanism of the formation of 3 was studied using model fermentation and model sulfuration media spiked with 1 or 2 as potential precursors. Possible degradation products such as kynurenamine (4) and kynurenic acid (5), or skatole (6), 2-oxoskatole (7), 2-formamidoacetophenone (8), 2-oxindole-3-acetic acid (9), and 3-(2-formylaminophenyl)-3-oxopropionic acid (10) were evaluated by HPLC-UV of the fermentation and sulfuration media and comparison with synthesized 7, 8, 9, and 10. The synthesis of the possible precursor 4-(2-aminophenyl)-2,4-dioxobutanoic acid (11), a proposed metabolite of 1 failed because a spontaneous cyclization yields 5 and N-oxo-kynurenic acid (12), but not 11. It could be shown that the formation of 3 is triggered by an oxidative degradation of 2 after sulfuration with potassium bisulfite via the intermediates 10 and 8. However, no formation of 3 occurred during sulfuration of a model wine spiked with 1 or during fermentation of a model must spiked with 1 or 2.

Acetophenones↗

Effect of fungicide residues on the aromatic composition of white wine inoculated with three Saccharomyces cerevisiae strains.

The effects of three fungicide residues (cyprodinil, fludioxonil, and pyrimethanil) on the aromatic composition (acids, alcohols, and esters) of Vitis vinifera white wines (var. Airén) inoculated with three Saccharomyces cerevisiae strains (syn. bayanus, cerevisiae, and syn. uvarum) are studied. The aromatic exponents were extracted and concentrated by adsorption-thermal desorption and were determined by gas chromatography using a mass selective detector. The addition of the three fungicides at different doses (1 and 5 mg/L) produces significant differences in the acidic fraction of the aroma, especially in the assays inoculated with S. cerevisiae, although the final contents do not exceed the perception thresholds. The lower quality wines, according to isomeric alcohol content [(Z)-3-hexen-1-ol and 3-(methylthio)propan-1-ol] are those obtained by inoculation with S. cerevisiae(syn. bayanus) and addition of cyprodinil. The addition of fungicides in the assays inoculated with S. cerevisiae (syn. bayanus) produces an increase in the ethyl acetate and isoamyl acetate contents, which causes a decrease in the sensorial quality of the wine obtained.

Acids↗

Characterization of phenolic profiles of Northern European berries by capillary electrophoresis and determination of their antioxidant activity.

Berries are known to contain phenolic substances (i.e., flavonoids and phenolic acids), which comprise two large and heterogeneous groups of biologically active nonnutrients. This investigation evaluated the content and profile of the phenolic compounds present in six different berries found in Northern Europe. The latter included bilberry (Vaccinium myrtillus), cowberry (Vaccinium vitis-idaea), cranberry (Vaccinium oxycoccus), strawberry (Fragaria ananassa), black currant (Ribes nigrum), and red currant (Ribes rubrum). The study was focused on two areas. The first involved the extraction and analysis of berries for total phenolic content and determination of their antioxidant activity. The total phenolic level of berries was correlated with their antioxidant activity. Second, the berry extracts were analyzed by capillary electrophoresis to determine the content and profile of selected bioactive compounds. The analytes of interest included trans-resveratrol, cinnamic acid, ferulic acid, p-coumaric acid, quercetin, and morin.

Antioxidants↗

Cutin composition of five finnish berries.

The raw cutin (i.e., extractive-free isolated cuticular membrane) fraction from Finnish berries, sea buckthorn (Hippophaë rhamnoides), black currant (Ribes nigrum), cranberry (Vaccinium oxycoccos), lingonberry (Vaccinium vitis-idaea), and bilberry (Vaccinium myrtillus), was depolymerized by NaOMe-catalyzed methanolysis. The composition of cutin monomers was determined by GC-(EI)MS analysis either as methyl esters or as TMSi esters, with OH groups derivatized to TMSi ethers. There was a notable difference in the degree of depolymerization, ranging from 6 to 47%. The extractive-free berry cuticle, that is, raw cutin, thus contains <50% polyester polymer cutin. The predominant cutin monomers were C(16) and C(18) omega-hydroxy acids with midchain functionalities, mainly epoxy and hydroxyl groups. Typically, the major compounds were 9,10-epoxy-18-hydroxyoctadecanoic acid, 10,16-dihydroxyhexadecanoic acid, 9,10,18-trihydroxyoctadecanoic acid, 9,10-epoxy-18-hydroxyoctadec-12-enoic acid, and 18-hydroxyoctadec-9-enoic acid. The amount of epoxyacids was rather high in sea buckthorn ( approximately 70%) and cranberry ( approximately 60%), compared with the other berries. The black currant cutin differed from that of the other berries with a significant portion of hydroxyoxohexadecanoic acid ( approximately 12% of total monomers). This investigation of the cuticular hydroxy acids of five Finnish berries is part of the exploitation of the northern natural resources related to the chemical composition, nutritional value, and sensory properties.

Finland↗

Relation between Fatty Acid Content and Its Evolution during Fermentation and Utilization of Free Amino Acids in Vacuum-Filtered Viura Must.

The activity of proteins that transport amino acids changes depending on the unsaturated residue which is introduced into the phospholipids of Saccharomycescerevisiae membrane. The aim of this study was to observe the influence of fatty acid content and its evolution during fermentation on the utilization of free amino acids in Vitis vinifera var. Viura must clarified by vacuum filtration. The results demonstrate that filtration of the must did not reduce the concentration of free amino acids, but it did reduce fatty acids, especially the unsaturateds. In the first half of fermentation (up to 50% sugar consumption), fatty acid utilization was different in each sample without involving an alteration in the uptake of amino acids. In the second half of fermentation (from 50% sugar consumption until the end of fermentation), the clarified sample had greater consumption of saturated fatty acids than unsaturated, in contrast to the control; this, along with other factors, produced a greater liberation of free amino acids in the filtered sample.

Journal Article↗

Regional origin assignment of red wines from Valencia (Spain) by (2)H NMR and (13)C IRMS stable isotope analysis of fermentative ethanol.

The use of the stable hydrogen and carbon isotope ratios of fermentative ethanol as suitable environmental fingerprints for the regional origin identification of red wines from Valencia (Spain) has been explored. Monovarietal Vitis vinifera L. cvs. Bobal, Tempranillo, and Monastrell wines have been investigated by (2)H NMR and (13)C IRMS for the natural ranges of site-specific (2)H/(1)H ratios and global delta(13)C values of ethanol over three vintage years. Statistically significant interregional and interannual (2)H and (13)C abundance differences have been noticed, which are interpreted in terms of environmental and ecophysiological factors of isotope content variation. Multivariate discriminant analysis is shown to provide a convenient means for integration of the classifying information, high discriminating abilities being demonstrated for the (2)H and (13)C fingerprints of ethanol. Reasonable differentiation results are achieved at a microregional scale in terms of geographic provenance and even grapevine genotypic features.

Carbon Isotopes↗

Effect of pesticide residues on the aromatic composition of red wines.

The influence of pesticide residues on the aromatic composition (major and minor volatiles) of red wines made from Vitis vinifera (Monastrell var.) was studied by comparing the concentration of aromas in wines made from grapes subjected (or not) to phytosanitary treatment with chlorpyrifos, fenarimol, mancozeb, metalaxyl, penconazole, and vinclozolin, according to the agricultural practice of the area. The analytical determination of the major volatiles was made by gas chromatography using a flame ionization detector, while the minor volatiles were determined by adsorption-thermal desorption gas chromatography using a mass selective detector. There were significant differences between the ethyl acetate, methanol, isobutanol, and diethylacetal levels of the control wine and that containing chlorpyrifos residues, although only the ethyl acetate exceeded the olfactory threshold. With regard to the minor volatiles, significant differences were detected in the concentrations of some esters, aldehydes, and acids. However, only isoamyl acetate exceeded the olfactory threshold in wines containing residues of chlorpyrifos, fenarimol, and vinclozolin.

Chlorpyrifos↗

Dimerization of resveratrol by the grapevine pathogen Botrytis cinerea.

Resveratrol (trans-3,4',5-trihydroxystilbene) is produced by grapes (Vitis spp.) in response to microbial attack by the fungal grapevine pathogen Botrytis cinerea. Several reports indicate that pathogenic B.cinerea strains are capable of biotransforming resveratrol into an assortment of unidentified oxidized metabolites as a means of reducing the antifungal effects of resveratrol and facilitating Botrytis invasion into host-plant tissues. Studies utilizing growing incubations of Botrytis cinerea ATCC 11542 with resveratrol resulted in the production of three new (restrytisols A-C) (1-3) and three known (resveratrol trans-dehydrodimer, leachinol F, and pallidol) oxidized resveratrol dimers. All of the metabolites were evaluated for their anti-HIV-1, cytotoxic, and cyclooxygenase (COX) I and COX II inhibitory activities.

Animals↗

Qualitative and quantitative variation among volatile profiles induced by Tetranychus urticae feeding on plants from various families.

Many plant species are known to emit herbivore-induced volatiles in response to herbivory. The spider mite Tetranychus urticae Koch is a generalist that can feed on several hundreds of host plant species. Volatiles emitted by T. urticae-infested plants of 11 species were compared: soybean (Glycine max), golden chain (Laburnum anagyroides), black locust (Robinia pseudo-acacia), cowpea (Vigna unguiculata), tobacco (Nicotiana tabacum), eggplant (Solanum melalonga), thorn apple (Datura stramonium), sweet pepper (Capsicum annuum), hop (Humulus lupulus), grapevine (Vitis vinifera), and ginkgo (Ginkgo biloba). The degree to which the plant species produced novel compounds was analyzed when compared to the odors of mechanically damaged leaves. Almost all of the investigated plant species produced novel compounds that dominated the volatile blend, such as methyl salicylate, terpenes, oximes, and nitriles. Only spider mite-infested eggplant and tobacco emitted a blend that was merely quantitatively different from the blend emitted by mechanically damaged or clean leaves. We hypothesized that plant species with a low degree of direct defense would produce more novel compounds. However, although plant species with a low direct defense level do use indirect defense to defend themselves, they do not always emit novel compounds. Plant species with a high level of direct defense seem to invest in the production of novel compounds. When plant species of the Fabaceae were compared to plant species of the Solanaceae, qualitative differences in spider mite-induced volatile blends seemed to be more prominent in the Fabaceae than in the Solanaceae.

Animals↗

[Efficacy of the red vine leaf extract AS 195 in Chronic Venous Insufficiency].

An open, observational clinical study to investigate the onset of action of the extract of the red vine-leaf AS 195 has been conducted in 39 patients suffering from Chronic Venous Insufficiency [CVI; grade I or II according to the Widmer classification (grade 2 to 4 of the international CEAP classification)]. Symptoms of CVI in the patients included oedema and subjective clinical complaints. The patients received 180 mg of extract twice daily (360 mg in total) during six weeks. The parameters investigated were objective measurements (lower leg volume by water plethysmography and the circumference of the leg at a fixed height), as well as subjective criteria such as heaviness and pain in the leg, measured using visual analogue scales (VAS). A clear and significant effect was already observed after two weeks of treatment. This effect was still present and increased slightly after four and six weeks in all objective and subjective parameters tested in this study. The decrease of the calf volume was statistically significant after 2, 4 and 6 weeks. The calf volume decreased on average by 26.3 ml after 2 weeks (t=-5.37 p<0.001), 29.8 ml after 4 weeks (t=-6.44 p<0.001) and 32.7 ml after 6 weeks (t=-7.67<0.001). The intensity of the sensation of heaviness and pain decreased also statistically significantly after 2, 4 and 6 weeks. The measured intensity on the VAS decreased on the average by 2.2 points after 2 weeks (t=-8.21 p<0.001), 2.6 points after 4 weeks (t=-10.93 p<0.001) and 2.6 points after 6 weeks (t=-10.75 p<0.001). In general, the treatments were well tolerated. Three patients prematurely stopped the trial for reasons not linked to the treatment or to the disease. This study has shown a fast onset of action and an excellent efficacy of the vitis viniferae Folia AS 195 extract in Chronic Venous Insufficiency.

Adolescent↗

Steady-state chlorophyll fluorescence (Fs) measurements as a tool to follow variations of net CO2 assimilation and stomatal conductance during water-stress in C3 plants.

Water stress experiments were performed with grapevines (Vitis vinifera L.) and other C3 plants in the field, in potted plants in the laboratory, and with detached leaves. It was found that, in all cases, the ratio of steady state chlorophyll fluorescence (Fs) normalized to dark-adapted intrinsic fluorescence (Fo) inversely correlated with non-photochemical quenching (NPQ). Also, at high irradiance, the ratio Fs/Fo was positively correlated with CO2 assimilation in air, with electron transport rate calculated from fluorescence, and with stomatal conductance, but no clear correlation was observed with qP. The significance of these relationships is discussed. The ratio Fs/Fo, measured with a portable instrument (PAM-2000) or with a remote sensing FIPAM system, provides a good method for the early detection of water stress, and may become a useful guide to irrigation requirements.

Journal Article↗