UHPLC-Q-Orbitrap /MS2 identification of (+)-Catechin oxidation reaction dimeric products in red wines and grape seed extracts.

B-type procyanidin dimers and (+)-catechin dimeric oxidation products were analyzed in grape seed extracts and red wines (UHPLC-Q-Orbitrap MS). The different dimers had different fragmentation patterns according to their interflavan linkage position. Oxidation dimeric compounds had a specific fragment ion at m/z 393, missing for B-Type dimers fragmentations. A fragment ion at m/z 291 occurred and was specific for oxidation dimeric compounds with a COC linkage. Higher level oxidation products had abundant specific fragments: m/z 425, 397 and 245. These fragmentations were useful to identify them in complex samples such as grape seed extracts and wines. Three grape varieties and three ripening stages were selected and the corresponding seed extracts were obtained. The analyses revealed an increasing trend for the oxidation markers during grape ripening. The analysis of Syrah wines (2018, 2014, 2010) showed a decreasing trend of these molecules during wine ageing which might be due to further oxidation.

A modified data filtering strategy for targeted characterization of polymers in complex matrixes using drift tube ion mobility-mass spectrometry: Application to analysis of procyanidins in the grape seed extracts.

BACKGROUNDS: Polymers, widely existing in food or dietary materials, have been attracting researchers, facing challenges, and needing effective strategies on targeted characterization in complex matrixes. METHODS: A modified data filtering strategy (including locating with drift time and m/z ranges, multiple mass defect filtering, validating MS information, and evaluating MS/MS spectra) was developed and applied for procyanidins in the grape seed extracts (GSE) using drift tube ion mobility-mass spectrometry. The procyanidin ions' trendlines were predicted by multi-model regression. Their collision cross-sections (CCSs) were calculated using single-field methods. RESULTS AND DISCUSSION: Totally, 769 CCSs belonging to 686 procyanidins with polymer degrees at 1-15 were characterized. The exponent regression was the most reasonable model (r2 ≥ 0.9379) to reveal the trendlines. The change tendency of CCSs with their polymer degrees, charge states, and linkage types were investigated. CONCLUSION: This study provided an innovative strategy for targeted characterization of polymers in complex matrixes.

Natural extracts from grape seed and stem by-products in combination with colloidal silver as alternative preservatives to SO2 for white wines: Effects on chemical composition and sensorial properties.

The search for alternative additives to sulfur dioxide, with antioxidant and antimicrobial activity, in the production of wines is one of the current objectives of the enological industry. In the present study, aqueous extracts obtained from winery byproducts (grape seeds and stems), alone or in combination with colloidal silver complex, have been used in white vinification. The antimicrobial effect of the extracts was similar to that of sulfur dioxide, being more effective on lactic and acetic bacteria in those wines to which colloidal silver was added. The effect on the color, the phenolic compounds and the volatile fraction of the wines was evaluated, as well as their sensory profile. The use of both extracts modified the color of the wines, increasing the chromatic parameters a* and b*, indicating a browning tendency, although no other signs of oxidation were found. Wines with seed extracts contained higher amounts of flavan-3-ols, and a significant increase in some volatile compounds such as fatty acid ethyl esters and benzene compounds, which were identified in the extracts. From a sensorial point of view, the wines with stem extracts were the most similar to those elaborated with SO2, detecting a certain bitterness in wines with seed extracts.

The impact of distillation process on the chemical composition and potential prebiotic activity of different oligosaccharidic fractions extracted from grape seeds.

The study was designed to evaluate how the distillation process is able to affect the composition and bioactivity of oligosaccharides contained in the grape seeds. Different oligosaccharidic fractions have been extracted both before and after grape pomace distillation in order to valorize this by-product. A multistep solid-phase extraction approach (C-18 and carbograph cartridges) has been applied to purify and fractionate the oligosaccharidic compounds. Chemical characterization of the fractions was performed using a UPLC-ESI-MSn method. Complex oligosaccharides consist principally of neutral oligosaccharides rich in arabinose and glucose. Then, the oligosaccharides contained in the different fractions as potential functional ingredients with prebiotic activity toward well-known probiotic bacteria, such as Lactobacillus acidophilus and Lactobacillus plantarum, were evaluated. Data showed how, in some combination, oligosaccharidic fractions obtained may be considered a novel "functional ingredient" with potential prebiotic activity mainly towards L. acidophilus.

Characterization, anti-oxidative effect of grape seed powder and in silico affinity profiling of polyphenolic and extra-phenolic compounds for calpain inhibition.

Vitis vinifera grape is a highly cultivated crop and solid wastes generated by the wine industry are largely under exploited. Plentiful studies have intended analyzing the polyphenolic content of grape seeds but characterization of non phenolic compounds is rather scarce. The present study aimed at the selective extraction of lipid, phenolic and aqueous phases from grape seed powder (GSP) in order to establish their intimate composition, as well as their antioxidant and chelating properties underlying partly their biological effects. Major non phenolic compounds identified in the lipid phase were glyceryl-monostearate and 2-monostearin whereas fructofuranose and sucrose were the most abundant in the aqueous phase. Among the most abundant compounds detected in the various phases, the polyphenol quercetin exhibited the best affinity and free binding energy towards the active site of the calcium-dependent protease calpain. Polyphenols likely constitute the bioactive part of GSP that should be exploited as safe modulators of intracellular signaling which is likely at the basis of their health beneficial effects. Nevertheless other compounds as lipids or sugars should be valorized along with polyphenols to improve their bioavailability into highly protected organs as brain or eye.

Determination of Total Phenolic Content Using the Folin-C Assay: Single-Laboratory Validation, First Action 2017.13.

A single-laboratory validation of a method using Folin & Ciocalteu's phenol reagent (Folin-C reagent) for determination of total phenolic content of selected dietary supplement extracts was performed. The method is composed of a water extraction of dried extracts with sonication followed by reaction with the Folin-C reagent. The resulting colorimetric reaction is measured at 765 nm and compared with a standard curve generated with gallic acid standard solutions. The validation results were compared with Standard Method Performance Requirement (SMPR®) 2015.009, developed by the Stakeholder Panel on Dietary Supplements. The method demonstrated acceptable within-day RSDr of 1.96-7.47% for the five matrixes studied (grape seed extract, grape skin extract, black tea extract, green coffee extract, and cocoa extract). When gallic acid was spiked into maltodextrin (a surrogate dietary supplement carrier) at 30 or 70%, the recovery ranged from 91 to 104%, within the acceptable range established by SMPR 2015.009. Selectivity testing with glucose, fructose, and sucrose demonstrated no positive interference by these compounds. Finally, ruggedness studies demonstrated no significant effects due to changes in the heating apparatus, test material weight, read time after reaction, amount of Folin-C reagent, reaction time, reaction temperature, and amount of Na2CO3. The single-laboratory validation results support adoption of the method as First Action Official MethodSM 2017.13 and further evaluation in a collaborative study.

Optimization of Extraction of Hypoglycemic Ingredients from Grape Seeds and Evaluation of α-Glucosidase and α-Amylase Inhibitory Effects In Vitro.

Grape seeds full with oil and also rich in hypoglycemic ingredients. This study is to improve the comprehensive utilization of grape seeds and the quality of grape seeds oil. Extract water soluble active substances before extracting oil from grape seeds, which show the grape seeds in water soluble extracts containing TP, TC, PC contents, respectively, are 171.1 ± 1.1, 658.8 ± 0.2, 482.3 ± 0.2 (mg GSAE/g). Also we found GSAE samples containing catechin and epicatechin were 44.12 ± 0.21 mg/mL, 111.23 ± 1.29 mg/g, GSAE against α-glucosidase IC50 was 25.25 ± 0.53 g/mL and GSAE against α-amylase IC50 was 66.68 ± 1.1 g/mL, both were competitive way, the effects of Inhibitory were obviously better than that of acarbose. In addition, our process guarantee the fat ingredients remained in the grape seeds and not reduce functional components of oil, our study suggests that GSAE can be used to develop functional foods for prevention and treatment of diabetes and its complications. PRACTICAL APPLICATION: Grape seeds, by-products of brewing wines, are rich in polyphenol and other ingredients. The optimized extraction, composition of constituents and hypoglycemic activity were investigated in this study. The results showed that grape seeds were rich in anthocyanins and polyphenols and other active substances, inhibited α-glucosidase and α-amylase activity, which provide background and practical knowledge for the deep-processed products of grape seeds with high added value.

Edible Coating Using a Chitosan-Based Colloid Incorporating Grapefruit Seed Extract for Cherry Tomato Safety and Preservation.

Grapefruit seed extract (GSE)-containing chitosan-based coating was developed and applied to cherry tomatoes to protect them from Salmonella invasion and improve their storability. The coating colloids were produced by mixing a chitosan colloid (1% [w/w] chitosan) with GSE at various concentrations (0.5%, 0.7%, 1.0%, and 1.2% [w/w]) using high-shear mixing (10000 rpm, 2 min). Coatings with chitosan colloids containing GSE at 0.0%, 0.5%, 0.7%, and 1.0% (w/w) inactivated Salmonella on cherry tomatoes by 1.0 ± 0.3, 1.2 ± 0.3, 1.6 ± 0.1, and 2.0 ± 0.3 log CFU/cherry tomato, respectively. Coatings both with and without GSE (1.0%) effectively inhibited the growth of Salmonella and total mesophilic aerobes, reduced CO2 generation, and retarded titratable acidity decrease during storage at 10 and 25 °C. The advantage of incorporating GSE in the formulation was demonstrated by delayed microorganism growth and reduced weight loss at 25 °C. The chitosan-GSE coating did not affect lycopene concentration, color, and sensory properties (P > 0.05). Chitosan-GSE coating shows potential for improving the microbiological safety and storability of cherry tomatoes, with stronger efficacy at 25 °C than that of chitosan coating without GSE. PRACTICAL APPLICATION: A novel chitosan coating containing grape fruit seed extract (GSE) improved the microbiological safety against Salmonella and storability of cherry tomatoes without altering their flavor, demonstrating its strong potential as an effective postharvest technology. Chitosan coating containing GSE might be preferable over chitosan coating without GSE for application to tomatoes that are stored at room temperature in that it more effectively inhibits microbial growth and weight loss than the coating without GSE at 25 °C.

Phenolic content and antioxidant properties of seeds from different grape cultivars grown in Iran.

This study investigated the antioxidant activity and phenolic composition of seed extracts from three grape cultivars grown in Iran. Folin Ciocalteu method was used for the determination of the total phenolic contents and GC-MS was used for the analysis of phenolic compositions. 1,1-diphenyl-2-picrylhydrazyl (DPPH) method was used to evaluate the antioxidant activity. The highest and the lowest total phenolic contents of seed extract were found in the black and green grape, respectively. The content of individual phenols such as Frulic acid, Gentistic acid, Syringic acid, (+) Catechin, Chlorogenic acid and (-)- Epicatchin gallate was cultivars dependent. The antioxidant activity of the seed extracts ranged from 34.03% (Green) to 53.63% (Black). Generally, the Black grape seed extract with the total phenolic content (3 ± 0.01 mg tannic acid/g DM), DPPH (53.63 ± 0.34%), IC50 and AEAC (7.41 and 16.92 mg/mL) showed the highest level of total antioxidant capacity.

Protective effects of grape seed and skin extract against high-fat-diet-induced lipotoxicity in rat lung.

BACKGROUND: Obesity is a public health problem characterized by increased fat accumulation in different tissues. Obesity is directly linked to breathing problems and medical complications with lung, including obstructive sleep apnea syndrome, obesity hypoventilation syndrome, chronic obstructive pulmonary disease, asthma….In the present work, we aimed to investigate the effect of high fat diet (HFD) on lung lipotoxicity, oxidative stress, fatty acid composition and proportions in lung and implication in asthma development. The likely protection provided by grape seed extract (GSSE) was also investigated. METHODS: In order to assess HFD effect on lung and GSSE protection we used a rat model. We analyzed the lipid plasma profile, lung peroxidation and antioxidant activities (SOD, CAT and POD). We also analyzed transition metals (Ca2+, Mg2+, Zn2+ and iron) and lung free fatty acids using gas chromatography coupled to mass spectrometry (GC-MS). RESULTS: HFD induced lipid profile imbalance increasing cholesterol and VLDL-C. HFD also induced an oxidative stress assessed by elevated MDA level and the drop of antioxidant activities such as SOD, CAT and POD. Moreover, HFD induced mineral disturbances by decreasing magnesium level and increasing Calcium and iron levels. HFD induced also disturbances in lung fatty acid composition by increasing oleic, stearic and arachidonic acids. Interestingly, GSSE alleviated all these deleterious effects of HFD treatment. CONCLUSION: As a whole, GSSE had a significant preventive effect against HFD-induced obesity, and hence may be used as an anti-obesity agent, and a benefic agent with potential applications against damages in lung tissue.

Wine consumers' subjective responses to wine mouthfeel and understanding of wine body.

Wine mouthfeel is considered important for wine quality by experts, while consumers understanding of mouthfeel and the role of wine body in their wine choice is unknown. One experiment determined the influence of intrinsic wine mouthfeel on consumers' wine liking and emotions, and the other, how consumers understand the term wine body. The first experiment used a 2 astringency level×2 body level experimental design. The samples were base wine with; nothing added (control), added xanthan gum (for increased body), added grape seed extract (GSE, for increased astringency), and with both added xanthan gum and GSE. The consumer taste trial (n=112) indicated that wine with increased body did not influence wine liking and emotions; while increased astringency decreased liking and elicited more intense negative emotions. The second experiment examined consumers' knowledge of wine body through an online survey (n=136). Consumers described wine body most frequently using words such as flavour, fullness, and strength. Wine body was therefore understood by consumers predominantly as a holistic multi-sensory perception of flavour. Wine flavour was indicated by consumers to be the most important factor driving purchase decisions followed by balance of flavours and wine body. It is crucial that wine professionals carefully communicate wine characteristics to consumers to prevent possible misunderstandings such as the meaning of wine body and as a result better meet consumer expectations. In future, the term body may benefit from a clearer definition for academic research as well as industry.

Preparative high-speed counter-current chromatography separation of grape seed proanthocyanidins according to degree of polymerization.

Separation of plant proanthocyanidins remains a major challenge for scientists due to the structural diversity and complexity. In this work, a new and effective method was developed for preparative separation of grape seed proanthocyanidins (GSPs) according to degree of polymerization (DP) by high-speed counter-current chromatography (HSCCC). Under the optimized HSCCC conditions, GSPs could be separated into seven distinct fractions (F1-F7) with mean degree of polymerization in increasing order, from 1.44 to 6.95. High yields for these fractions (53.7, 12.2, 29.5, 30.2, 11.2, 50.8 and 169.8mg, respectively) were achieved by only one-step HSCCC of 400mg of GSPs. Further, seventeen individual proanthocyanidins, most of which are commercially not available, were efficiently isolated by re-chromatography on HSCCC or prep-HPLC; each of the isolated compounds presented high yields (7.1-78.9mg) and high purity (70.0-95.7%). The positive correlation was observed between DP and antioxidant activity of proanthocyanidins.

Evaluation of the influence of white grape seed extracts as copigment sources on the anthocyanin extraction from grape skins previously classified by near infrared hyperspectral tools.

Hyperspectral imaging has been used to classify red grapes (Vitis vinifera L.) according to their predicted extractable total anthocyanin content (i.e. extractable total anthocyanin content determined by a hyperspectral method). Low, medium and high levels of predicted extractable total anthocyanin content were established. Then, grape skins were split into three parts and each part was macerated into a different model wine solution for a three-day period. Wine model solutions were made up with different concentration of copigments coming from white grape seeds. Aqueous supernatants were analyzed by HPLC-DAD and extractable anthocyanin contents were obtained. Principal component analyses and analyses of variance were carried out with the aim of studying trends related to the extractable anthocyanin contents. Significant differences were found among grapes with different levels of predicted extractable anthocyanin contents. Moreover, no significant differences were found on the extractable anthocyanin contents using different copigment concentrations in grape skin macerations.

Antioxidative and prooxidative effects in food lipids and synergism with α-tocopherol of açaí seed extracts and grape rachis extracts.

Extracts of açaí seed and of grape rachis alone or in combination with α-tocopherol were evaluated as antioxidants in (i) bulk soybean oil, (ii) soybean oil liposomes and (iii) soybean-oil/water emulsions. The extracts made with 57% aqueous ethanol showed an antioxidant activity not dependent on concentration for grape rachis extracts and a concentration-dependent prooxidative activity for açaí seed extracts in bulk soybean oil. Both the extracts, however, protected liposome suspensions and oil/water emulsions against lipid oxidation. Synergism was demonstrated when extracts were combined with α-tocopherol, effects explained by the solubility of extract components in the water-phase and of α-tocopherol in the lipid-phase. Phenolic profiling of the extracts by U-HPLC-ESI-LTQ-MS was used to identify active antioxidants. Açaí seed and grape rachis extracts served as good sources of procyanidins and flavan-3-ols, imparted high antioxidant activity especially when combined with α-tocopherol and are suggested for protection of food oil/water emulsions.

Preparative HSCCC isolation of phloroglucinolysis products from grape seed polymeric proanthocyanidins as new powerful antioxidants.

Polymeric proanthocyanidins isolated from a grape seed phenolic extract were hydrolysed in the presence of phloroglucinol into monomer catechins and their nucleophile derivatives. Each of the phloroglucinolysis products was successfully separated and isolated in large amount by semi-preparative HSCCC technique under the optimized conditions based on a selection of suitable solvent system. The optimized solvent system consisted of n-hexane-ethyl acetate-water (1:80:80, v/v/v) with a combination of head-tail and tail-head elution modes. By only one-step HSCCC separation, the purity of each obtained phloroglucinolysis product, including monomer catechins and their nucleophile derivatives was above 76%, verified by UPLC. The structures of these products were tentatively identified by UPLC based on their retention time and further confirmed by MS and (1)H NMR analysis. Furthermore, by DPPH, ABTS and FRAP assays, it was verified that all these phloroglucinolysis products possessed strong antioxidant activities, being catechin-nucleophile derivatives more powerful than free catechins.

Effects of natural antioxidants on colour stability, lipid oxidation and metmyoglobin reducing activity in raw beef patties.

BACKGROUND: Minced meats undergo oxidative changes and develop rancidity more quickly than intact muscle since grinding exposes more of the muscle surface to air and microbial contamination. Due to concerns about toxicological safety of synthetic antioxidants, recent studies have put more focus on natural antioxidant compounds derived from food components. METHODS: The effects of four natural antioxidants (vitamin E, carnosine, grape seed extract and tea catechins) on oxidative processes and metmyoglobin reducing activity in raw beef patties during refrigerated (4°C) storage were investigated and the results were compared with butylated hydroxyanisole treatment patties. The correlation of lipid oxidation, colour and metmyoglobin reducing activity of beef patties were also studied. RESULTS: Samples treated with carnosine had the highest redness values on the eighth day. Tea catechins, vitamin E and grape seed extract showed higher protective effect against lipid oxidation than carnosine. Metmyoglobin reducing activity increased greatly in all samples during the storage. Significant correlation between redness value and lipid oxidation was demonstrated, while a weak correlation between metmyoglobin reducing activity and any other parameters was shown.

Low-fat beef patties with augmented omega-3 fatty acid and CLA levels and influence of grape seed extract.

The effects of raising the omega-3 fatty acid (FA), conjugated linoleic acid (CLA), or omega-3 FA plus CLA levels on beef by means of dietary supplementation and of adding grape seed extract (250 mg/kg meat product) in beef patties stored at 2 ± 1 °C in aerobic packaging under simulated retail display conditions for 6 d was evaluated by measuring the thiobarbituric acid reactive substances (TBARS), pH, and instrumental color measurement values and by means of sensory analysis. The pH, instrumental color measurements, and sensory attribute values for patties made from beef with augmented omega-3 FA and/or CLA contents were similar to the values for the control patties made from beef from animals fed a conventional diet. Adding GSE lowered oxidation levels on day 6 (P < 0.001) and did not affect the instrumental color or sensory analysis results during the display period. This suggests that omega-3 FA and CLA-augmented beef could be used to make low-fat beef patties having characteristics similar to those of conventional beef patties while being more in keeping with currently recommended nutritional guidelines.

Gallic acid is the major component of grape seed extract that inhibits amyloid fibril formation.

Many protein misfolding diseases, for example, Alzheimer's, Parkinson's and Huntington's, are characterised by the accumulation of protein aggregates in an amyloid fibrillar form. Natural products which inhibit fibril formation are a promising avenue to explore as therapeutics for the treatment of these diseases. In this study we have shown, using in vitro thioflavin T assays and transmission electron microscopy, that grape seed extract inhibits fibril formation of kappa-casein (κ-CN), a milk protein which forms amyloid fibrils spontaneously under physiological conditions. Among the components of grape seed extract, gallic acid was the most active component at inhibiting κ-CN fibril formation, by stabilizing κ-CN to prevent its aggregation. Concomitantly, gallic acid significantly reduced the toxicity of κ-CN to pheochromocytoma12 cells. Furthermore, gallic acid effectively inhibited fibril formation by the amyloid-beta peptide, the putative causative agent in Alzheimer's disease. It is concluded that the gallate moiety has the fibril-inhibitory activity.

Delivery of natural polyphenols by polymeric nanoparticles improves the resistance of endothelial progenitor cells to oxidative stress.

PURPOSE: Bone marrow-derived endothelial progenitor cells (EPCs) circulate into peripheral blood and significantly contribute to neo-vascularisation and re-endothelialisation as part of the process of vascular repair. Several studies have reported decreased EPC number in the presence of oxidative stress. Aim of this study was to evaluate the validity of mucoadhesive polymeric nanoparticles as a delivery system of natural products able to protect EPCs from oxidative stress. METHODS: The total polyphenol content and antioxidant capacity of red grape seed extract (GSE) either pre-veraison (p-GSE) or ripe (r-GSE) were measured. Cell viability was evaluated by WST-1 assay. Nanoparticles were prepared by ionotropic crosslinking of two structurally different thiolated quaternary ammonium-chitosan conjugates. A hyaluronic acid solution, containing p-GSE or r-GSE, was added to a stirred solution of each of the two chitosan derivatives to obtain p- or r-GSE loaded nanoparticles (NP) of two types. RESULTS: Both GSE types demonstrated strong antioxidant capacity. p-GSE showed a higher content in total polyphenols compared to r-GSE. NP size was in the 310-340 nm range, with 24 h stability, and nearly 100% encapsulation efficiency for both GSE types. NP were internalized by cells to an extent related directly with their surface charge intensity. GSE-NP uptake significantly improved cell viability and resistance to oxidation. CONCLUSIONS: Nanotechnology has a great potential in nutraceutical delivery. The present results suggest that NP is a highly promising polyphenol carrier system particularly useful to protect EPCs from oxidative stress, thus improving their survival.