Glucose and fructose levels on grape skin: interference in Lobesia botrana behaviour

The vineyard produces one of the most important crops in Portugal and is affected by various pests, such as the Lobesia botrana moth, which can be attracted chemically by compounds on the grape skin.In this study, the sugar contents (glucose and fructose) on grape skin were analysed for five Portuguese Vitis vinifera grape varieties and compared in grape juice and whole grapes. Samples were harvested in July, August and September of 2001 and 2002.The quantification was attained employing a new methodology, which comprised the preparation of a methanol solution to extract the compounds from the grape skin and further chromatographic analyses by liquid chromatography (LC) with refractive index (RI) detection. The samples designated by whole grapes were homogenised, centrifuged and filtrated, while the juice was obtained by crushing and filtration.Grape skin has shown higher levels of glucose than fructose, but both compounds increased from green berry (July) to veraison (August) and had a slight decrease from veraison to harvest (September).As expected, the glucose and fructose contents (total sugar) were higher in the whole grape than in the juice and much higher than in the grape skin itself. For Loureiro and Trajadura varieties, the levels of both sugars on grape skin were very similar. This resemblance strengthened the statement of the decisive role of this parameter in female L. botrana oviposition behaviour.     

Rapidly differentiating grape seeds from different sources based on characteristic fingerprints using direct analysis in real time coupled with time‐of‐flight mass spectrometry combined with chemometrics

The seeds of grapevine (Vitis vinifera) are a byproduct of wine production. To examine the potential value of grape seeds, grape seeds from seven sources were subjected to fingerprinting using direct analysis in real time coupled with time‐of‐flight mass spectrometry combined with chemometrics. Firstly, we listed all reported components (56 components) from grape seeds and calculated the precise m/z values of the deprotonated ions [M–H]^–. Secondly, the experimental conditions were systematically optimized based on the peak areas of total ion chromatograms of the samples. Thirdly, the seven grape seed samples were examined using the optimized method. Information about 20 grape seed components was utilized to represent characteristic fingerprints. Finally, hierarchical clustering analysis and principal component analysis were performed to analyze the data. Grape seeds from seven different sources were classified into two clusters; hierarchical clustering analysis and principal component analysis yielded similar results. The results of this study lay the foundation for appropriate utilization and exploitation of grape seed samples. Due to the absence of complicated sample preparation methods and chromatographic separation, the method developed in this study represents one of the simplest and least time‐consuming methods for grape seed fingerprinting.     

Health Beneficial Properties of Grapevine Seed Extract and Its Influence on Selected Biochemical Markers in the Blood,Liver and Kidneys of Rattus norvegicus

Cadmium (Cd) is a heavy metal that occurs in all areas of the environment, including the food chain. In the body, it causes oxidative stress by producing free radicals that are harmful to the cells. Grape seed extract (GSE) contains a wide range of biologically active components that help to neutralize the adverse effects of free radicals. In this study, the effects of GSE prepared form semi-resistant grapevine cultivar Cerason, which is rich in phenolics, on biochemical markers of brown rats exposed to the effects of cadmium were monitored. GSE increased the plasma antioxidant activity and, in the kidneys and the liver, Cd content was significantly lowered by GSE co-administration. Accordingly, the increase in creatinine content and alanine aminotransferase activity and the decrease of catalase and superoxide dismutase activities caused by cadmium were slowed down by GSE co-administration. The results of this work reveal that grape seed extract offers a protective effect against the intake of heavy metals into the organism.     

Biotransformation from geraniol to nerol by immobilized grapevine cells (V. vinifera)

The abilities of two grapevine cell suspensions (Vitis vinifera L. cv. Gamay Fréaux andVitis vinifera L. cv. Monastrell) to biotransform geraniol into nerol in a biphasic system based on the culture medium and Miglyol 812 were compared. The Gamay grape cell suspension was able to transform higher concentrations of geraniol into nerol than the Monastrell one. Gamay grape cells were immobilized in both calcium alginate beads and polyurethane foams. The cytotoxic effect of increasing concentrations of geraniol, as well as the ability of the immobilized cells to biotransform geraniol into nerol, was checked. Immobilization proved to be advantageous in protecting cells against the toxicity of the substrate. Furthermore, immobilization also seemed to have an effect on the secondary metabolism, the cells immobilized in polyurethane foams being more efficient at performing the isomerization process (40% conversion of geraniol into nerol) than both the freely suspended and calcium alginate immobilized cells (20% conversion).     

Rapid monitoring of grapevine reserves using ATR–FT-IR and chemometrics

Predictions of grapevine yield and the management of sugar accumulation and secondary metabolite production during berry ripening may be improved by monitoring nitrogen and starch reserves in the perennial parts of the vine. The standard method for determining nitrogen concentration in plant tissue is by combustion analysis, while enzymatic hydrolysis followed by glucose quantification is commonly used for starch. Attenuated total reflectance Fourier transform infrared spectroscopy (ATR–FT-IR) combined with chemometric modelling offers a rapid means for the determination of a range of analytes in powdered or ground samples. ATR–FT-IR offers significant advantages over combustion or enzymatic analysis of samples due to the simplicity of instrument operation, reproducibility and speed of data collection. In the present investigation, 1880 root and wood samples were collected from Shiraz, Semillon and Riesling vineyards in Australia and Germany. Nitrogen and starch concentrations were determined using standard analytical methods, and ATR–FT-IR spectra collected for each sample using a Bruker Alpha instrument. Samples were randomly assigned to either calibration or test data sets representing two thirds and one third of the samples respectively. Signal preprocessing included extended multiplicative scatter correction for water and carbon dioxide vapour, standard normal variate scaling with second derivative and variable selection prior to regression. Excellent predictive models for percent dry weight (DW) of nitrogen (range: 0.10–2.65% DW, median: 0.45% DW) and starch (range: 0.25–42.82% DW, median: 7.77% DW) using partial least squares (PLS) or support vector machine (SVM) analysis for linear and nonlinear regression respectively, were constructed and cross validated with low root mean square errors of prediction (RMSEP). Calibrations employing SVM-regression provided the optimum predictive models for nitrogen (R² = 0.98 and RMSEP = 0.07% DW) compared to PLS regression (R² = 0.97 and RMSEP = 0.08% DW). The best predictive models for starch was obtained using PLS regression (R² = 0.95 and RSMEP = 1.43% DW) compared to SVR (R² = 0.95; RMSEP = 1.56% DW). The RMSEP for both nitrogen and starch is below the reported seasonal flux for these analytes in Vitis vinifera. Nitrogen and starch concentrations in grapevine tissues can thus be accurately determined using ATR–FT-IR, providing a rapid method for monitoring vine reserve status under commercial grape production.     

A Potential Valorization Strategy of Wine Industry by-Products and Their Application in Cosmetics—Case Study: Grape Pomace and Grapeseed

Grape pomace and grapeseed are agro-industrial by-products, whose inadequate treatment generates socioeconomic and environmental concerns. Nevertheless, it is possible to valorize them by extracting their bioactive compounds, such as antioxidants (phenolic compounds), vitamin E and fatty acids. The bioactive compounds were extracted using solid-liquid extraction. The yields for phenolic compounds were 18.4 ± 0.4% for grape pomace, and 17.4 ± 0.4%, for grapeseed. For the oil, the yields were 13.3 ± 0.2% and 14.5 ± 0.3% for grape pomace and grapeseed. Antioxidant capacity was assessed by the assay with 2,2-diphenyl-1-picrylhydrazyl (DPPH), and showed that phenolic extract has higher antioxidant capacity than the oils. Grape pomace and grapeseed extracts exhibit, correspondingly, values of 90.8 ± 0.8 and 87.5 ± 0.5 of DPPH inhibition and IC₅₀ of 48.9 ± 0.5 and 55.9 ± 0.7 μg_extract·mL_DPPH⁻¹. The antimicrobial capacity was assessed by the disk diffusion test, and revealed that, phenolic extracts inhibit the growth of Staphylococcus aureus and Staphylococcus epidermidis. The obtained extracts were incorporated in 10 face cream formulations, with slight modifications in quantities of formulation stabilizers. Their stability was studied for 35 days, and this revealed the possibility of incorporating extracts and oils obtained from by-products as antioxidants in cosmetics, and replacing synthetic ones. As a future recommendation, microencapsulation of the extracts should be performed, in order to increase their stability.     

A critical review on extraction techniques and gas chromatography based determination of grapevine derived sesquiterpenes

Vitis vinifera L. (common grapevine) is considered one of the major world fruit crops based on the extent of cultivated land and on its economic value. Grapevine is composed by several different chemical compounds, including sesquiterpenic ones. Sesquiterpenic compounds play a significant role in varietal aromas, contributing to fruity and flowery odours, and are also well known for their potential health benefits. The advantages and drawbacks of different extraction and gas chromatographic techniques used for the determination of sesquiterpenic compounds are critically presented. Also, the future trends of sample preparation and gas chromatographic tools are discussed. This review provides the state-of-the-art and the technical know-how for the researchers who want to start studying sesquiterpenic compounds on complex matrices such as grapevine products.     

Large-Scale Reassessment of In-Vineyard Smoke-Taint Grapevine Protection Strategies and the Development of Predictive Off-Vine Models

Smoke taint in wine is thought to be caused by smoke-derived volatile phenols (VPs) that are absorbed into grape tissues, trapped as conjugates that are imperceptible by smell, and subsequently released into wines as their free odor-active forms via metabolism by yeasts during fermentation. Blocking VP uptake into grapes would, therefore, be an effective way for vineyards to protect ripening grape crops exposed to smoke. Here, we re-evaluated a biofilm that had previously shown promise in pilot studies in reducing levels of smoke-derived VPs. A suite of nine free and acid-labile VPs were quantitated in Pinot Noir grapes that had been exposed to smoke after being coated with the biofilm one, seven or 14 days earlier. In contrast with earlier studies, our results demonstrated that in all cases, the biofilm treatments led to increased concentrations of both free and total VPs in smoke-exposed grapes, with earlier applications elevating concentrations of some VPs more than the later time points. Tracking VP concentrations through the grape ripening process demonstrated that some (phenol, p/m-cresol, and guaiacol) were not entirely sequestered in grapes as acid-labile conjugates, suggesting the presence of VP storage forms beyond simple glycosides. Free VPs in grapes, though a minor portion of the total, most clearly correlated with concentrations present in the resulting wines. Finally, red table grapes, available year round, were observed to replicate the effects of the biofilm treatments and were capable of transforming most VPs into acid-labile conjugates in under 24 h, indicating that they might be an effective model for rapidly assessing smoke-taint prophylactic products in the laboratory.     

Thinking Inside the Box: A Novel Approach to Smoke Taint Mitigation Trials

When bushfires occur near wine regions, grapevine exposure to smoke can taint grapes due to the uptake of smoke-derived volatile compounds that can subsequently impart unpleasant smoky, medicinal, burnt rubber and ashy characters to wine. Whereas early research sought to understand the effects of smoke on grapevine physiology, and grape and wine chemistry, research efforts have shifted towards the strategic imperative for effective mitigation strategies. This study evaluated the extent to which excised grape bunches could be reproducibly tainted during smoke exposure in a purpose-built ‘smoke box’. The volatile phenol composition of grapes exposed to smoke for 30 min was similar to that of smoke-affected grapes from field trials involving grapevine exposure to smoke. Some variation was observed between replicate smoke treatments, but implementing appropriate controls and experimental replication enabled the smoke box to be used to successfully evaluate the efficacy of several agrochemical sprays and protective coverings as methods for mitigating the smoke exposure of grapes. Whereas the agrochemical sprays did not provide effective protection from smoke, enclosing grape bunches in activated carbon fabric prevented the uptake of up to 98% of the smoke-derived volatile phenols observed in smoke-affected grapes. As such, the study demonstrated not only a convenient, efficient approach to smoke taint research that overcomes the constraints associated with vineyard-based field trials, but also a promising new strategy for preventing smoke taint.     

Production of Cellulolytic Enzymes by <Emphasis Type="Italic">Aspergillus phoenicis</Emphasis> in Grape Waste using Response Surface Methodology

The production of cellulolytic enzymes by the fungus Aspergillus phoenicis was investigated. Grape waste from the winemaking industry was chosen as the growth substrate among several agro-industrial byproducts. A 2 × 2 central composite design was performed, utilizing the amount of grape waste and peptone as independent variables. The fungus was cultivated in submerged fermentation at 30 °C and 120 rpm for 120 h, and the activities of total cellulases, endoglucanases, and β-glucosidases were measured. Total cellulases were positively influenced by the linear increase of peptone concentration and decrease at axial concentrations of grape waste and peptone. Maximum activity of endoglucanase was observed by a linear increase of both grape waste and peptone concentrations. Concentrations of grape waste between 5 and 15 g/L had a positive effect on the production of β-glucosidase; peptone had no significant effects. The optimum production of the three cellulolytic activities was observed at values near the central point. A. phoenicis has the potential for the production of cellulases utilizing grape waste as the growth substrate.     

Profiling monoterpenol glycoconjugation in Vitis vinifera L. cv. Muscat of Alexandria using a novel putative compound database approach,high resolution mass spectrometry and collision induced dissociation fragmentation analysis

In this work we present a novel approach for the identification of plant metabolites using ultrahigh performance liquid chromatography coupled to accurate mass time-of-flight mass spectrometry. The workflow involves developing an in-house compound database consisting of exact masses of previously identified as well as putative compounds. The database is used to screen accurate mass spectrometry (MS) data to identify possible compound matches. Subsequent tandem MS data is acquired for possible matches and used for structural elucidation. The methodology is applied to profile monoterpene glycosides in Vitis vinifera cv. Muscat of Alexandria grape berries over three developmental stages. Monoterpenes are a subclass of terpenes, the largest class of plant secondary metabolites, and are found in two major forms in the plant, “bound” to one or more sugar moieties or “free” of said sugar moieties. In the free form, monoterpenes are noted for their fragrance and play important roles in plant defense and as attractants for pollinators. However, glycoconjugation renders these compounds odorless, and it is this form that the plant uses for monoterpene storage. In order to gain insight into monoterpene biochemistry and their fate in the plant an analysis of intact glycosides is essential. Eighteen monoterpene glycosides were identified including a monoterpene trisaccharide glycoside, which is tentatively identified here for this first time in any plant. Additionally, while previous studies have identified monoterpene malonylated glucosides in other grapevine tissue, we tentatively identify them for the first time in grape berries. This analytical approach can be readily applied to other plants and the workflow approach can also be used for other classes of compounds. This approach, in general, provides researchers with data to support the identification of putative compounds, which is especially useful when no standard is available.     

Optimization of SPME-Arrow-GC/MS Method for Determination of Free and Bound Volatile Organic Compounds from Grape Skins

(1) Background: Solid phase microextraction (SPME)-Arrow is a new extraction technology recently employed in the analysis of volatiles in food materials. Grape volatile organic compounds (VOC) have a crucial role in the winemaking industry due to their sensory characteristics of wine.; (2) Methods: Box–Behnken experimental design and response surface methodology were used to optimise SPME-Arrow conditions (extraction temperature, incubation time, exposure time, desorption time). Analyzed VOCs were free VOCs directly from grape skins and bound VOCs released from grape skins by acid hydrolysis.; (3) Results: The most significant factors were extraction temperature and exposure time for both free and bound VOCs. For both factors, an increase in their values positively affected the extraction efficiency for almost all classes of VOCs. For free VOCs, the optimum extraction conditions are: extraction temperature 60 °C, incubation time 20 min, exposure time 49 min, and desorption time 7 min, while for the bound VOCs are: extraction temperature 60 °C, incubation time 20 min, exposure time 60 min, desorption time 7 min.; (4) Conclusions: Application of the optimized method provides a powerful tool in the analysis of major classes of volatile organic compounds from grape skins, which can be applied to a large number of samples.     

Per-O-methylation of neutral carbohydrates directly from aqueous samples for gas chromatography and mass spectrometry analysis

Per-O-methylation of neutral carbohydrates in one step by adding dimethyl sulfoxide, powdered sodium hydroxide, and methyl iodide directly to aqueous sample is described. The influence of the water on the methylation reaction is investigated. Solid powdered sodium hydroxide is very hygroscopic and can scavenge the water from sample if an additional excess of sodium hydroxide is added. The degree of per-O-methylation of carbohydrates is checked by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Gas chromatography-mass spectrometry analysis of mono- and disaccharides from grape juice is presented.     

Mass spectrometry in the study of anthocyanins and their derivatives: differentiation of Vitis vinifera and hybrid grapes by liquid chromatography/electrospray ionization mass spectrometry and tandem mass spectrometry

A mass spectrometric-based procedure for anthocyanin profiling was set up to distinguish authentic Vitis vinifera from hybrid red grapevine cultivars. 3-O-Monoglucoside and the related acetyl-, p-coumaryl- and caffeoyl-monoglucoside anthocyanins occurred only in Vitis vinifera, whereas 3,5-O-diglucoside and the substituted acetyl-, p-coumaryl-, feruloyl- and caffeoyl-diglucoside anthocyanins were the additional pigments in hybrid grapevines. The procedure was applied expressly to identify red grape cultivars based on the anthocyanin chemo-type determination. In particular, a red grape cultivar, having 3,5-O-diglucoside anthocyanins and a novel class of anthocyanin monoglucosides, such as cyanidin-3-O-, cyanidin-3-O-(6-O-acetyl)- and cyanidin-3-O-(6-O-p-coumaryl)pentoside, was classified as hybrid. A second vine cultivar, characterized exclusively by 3-O-monoglucoside anthocyanins, was included among the Vitis vinifera species. Anthocyanin profiling by mass spectrometry could represent the core of a chemotaxonomic procedure for distinguishing American and European grapevines based on the identification of post-synthetic anthocyanidin modification.     

trans-α-Necrodyl isobutyrate, the sex pheromone of the grape mealybug, Pseudococcus maritimus

Headspace volatiles of virgin female grape mealybugs contained a single compound that elicitited responses from antennae of males in GC-electroantennogram analyses. The structure, identified as (R^∗,R^∗)-trans-(3,4,5,5-tetramethylcyclopent-2-en-1-yl)methyl 2-methylpropanoate, an irregular, non-head-to-tail monoterpenoid, was confirmed by synthesis.     

Mapping Grape Berry Photosynthesis by Chlorophyll Fluorescence Imaging: The Effect of Saturating Pulse Intensity in Different Tissues

Grape berry development and ripening depends mainly on imported photosynthates from leaves, however, fruit photosynthesis may also contribute to the carbon economy of the fruit. In this study pulse amplitude modulated chlorophyll fluorescence imaging (imaging‐PAM) was used to assess photosynthetic properties of tissues of green grape berries. In particular, the effect of the saturation pulse (SP) intensity was investigated. A clear tissue‐specific distribution pattern of photosynthetic competence was observed. The exocarp revealed the highest photosynthetic capacity and the lowest susceptibility to photoinhibition, and the mesocarp exhibited very low fluorescence signals and photochemical competence. Remarkably, the seed outer integument revealed a photosynthetic ability similar to that of the exocarp. At a SP intensity of 5000 μmol m^?2 s^?1 several photochemical parameters were decreased, including maximum fluorescence in dark‐adapted (F_m) and light‐adapted (F'_m) samples and effective quantum yield of PSII (Φ_II), but the inner tissues were susceptible to a SP intensity as low as 3200 μmol m^?2 s^?1 under light‐adapted conditions, indicating a photoinhibitory interaction between SP and actinic light intensities and repetitive exposure to SP. These results open the way to further studies concerning the involvement of tissue‐specific photosynthesis in the highly compartmentalized production and accumulation of organic compounds during grape berry development.     

Grape extracts suppress the formation of preneoplastic foci and activity of fatty acid synthase in rat liver

This study was conducted to examine the effects of dietary grape extracts on preneoplastic foci formation in rat hepatocarcinogenesis, and related hepatic enzymes. Male Sprague-Dawley rats were fed basal diet or grape diet containing 15% concentrated grape extracts (68 bricks). The grape diet groups were divided into whole-period grape diet group (DEN-GW; grape diet group fed throughout experimental period) and postinitiation grape diet group (DEN-GP; grape diet group fed from post initiation stage) according to the starting time point of the grape diet. Hepatocarcinogenesis was induced by diethylnitrosamine (DEN; 200 mg/kg bw) and 2/3 partial hepatectomy (DEN-B; DEN-treated basal diet group, DEN-GW, and DEN-GP groups), while the control group treated with saline and sham operation (Control group). The formation of placental glutathione (GSH) S-transferase positive (GST-P+) foci in DEN-GW group was moderately but significantly suppressed, however, not in DEN- GP group. Thiobarbituric acid reactive substances content of DEN-GW group was significantly lower than that of DEN-B group. The activity of fatty acid synthase (FAS) in the grape diet groups was decreased about 1/2 of the DEN-B group. The content of GSH and GSH peroxidase activity were increased by carcinogen treatment, but not modulated by grape diet. The activities of GSH S-transferase, p-nitrophenol hydroxylase, and catalase were not affected by diet or treatment. Conclusively, the grape diet-induced reduction of FAS activity that was expressed highly in neoplastic tissues, might be one of the contributing mechanisms of hepatic cancer prevention.     

Biogeneration of C13-norisoprenoid compounds: experiments supportive for an apo-carotenoid pathway in grapevines

The first step of the proposed biogenetic pathway in grapes that leads from carotenoids to C₁₃-norisoprenoids involves the enzymatic degradation of carotenoids by regiospecific oxygenases. Chemical, photochemical and oxidase-coupled degradation of carotenoids to norisoprenoids have been studied in vitro and enzymatic systems shown to be involved in mammals. However, no enzymatic system has been shown to be involved in the formation of C₁₃-norisoprenoids in grapes, despite all recent studies carried out on grapevines supporting such a model. These findings include the preponderance of norisoprenoids possessing 13 carbon atoms that thereby indicates the specificity of cleavage, the configuration of the asymmetric centres and axes common to C₁₃-norisoprenoids and the corresponding carotenoids, the negative correlations observed between the levels of norisoprenoids and carotenoids during berry development, and the in vivo transfer of ¹³C markers from carotenoids to norisoprenoids in berries between véraison and berry maturity. All of these findings are major arguments in favour of the hypothesis that glycosylated C₁₃-norisoprenoids derive from carotenoids in grape berries. Carotenoids are mostly synthesised from the first stage of fruit formation until véraison, and then degrade between véraison and maturity to produce glycosylated C₁₃-norisoprenoids and other compounds.     

Development of hybrid elution systems for efficient purification of stilbenoids using centrifugal partition chromatography coupled to mass spectrometry

The phytochemical study of the root extract of the stilbenoid-rich Vitis riparia × Vitis berlandieri grapevine was carried out by centrifugal partition chromatography (CPC). For this reason, we developed a new elution mode we named back-step, which allowed us to obtain cleaner fractions and a more efficient separation process when used in conjunction with a classical elution approach. Three hydroxystilbenes: (E)-resveratrol, (E)-?-viniferin and (E)-vitisin C, with greater than 90% purity were thus obtained through such process, with minimal sample handling and purification steps. Online coupling of CPC to ESI mass spectrometry was used for optimization of the separation parameters and to facilitate the characterization of the stilbenoids. This study details the first phytochemical investigation of stilbenoids from the hybrid species together with a new elution mode able to widen the range of ARIZONA biphasic systems.     

Anthocyanin identification and composition of wild Vitis spp. accessions by using LC–MS and LC–NMR

The composition and concentration of anthocyanins of grape berry skins were analyzed in order to assess phenotypic variation between four grape wine varieties belonging to 4 different species: Vitis vinifera, Vitis amurensis, Vitis cinerea and Vitis X champinii. High-performance liquid chromatography coupled to mass spectrometry (LC–MS) and NMR spectroscopy (LC–NMR) were used to separate and identify the structure of anthocyanins present in these species. Combination of LC–MS and LC–NMR data resulted in the identification of 33 anthocyanins. In particular, newly reported cis isomers of p-coumaric-derivatives were identified (petunidin-, peonidin- and malvidin-3-(6-p-coumaroyl)-5-diglucoside). In V. cinerea and V. vinifera, anthocyanins were monoglucoside derivatives whereas in V. amurensis and V. X champinii, both mono- and diglucoside derivatives were identified. Malvidin-, delphinidin- and petunidin-derivatives were, respectively, the most abundant components in V. cinerea and V. vinifera, V. amurensis and V. X champinii.