Isolation of Hydrolyzable Tannins from Castanea sativa Using Centrifugal Partition Chromatography.

Castanea sativa wood is a rich source of hydrolyzable tannins, known for their diverse bioactivities. To investigate these bioactive properties further, it is crucial to isolate and characterize hydrophilic compounds effectively. To address this issue, we developed a centrifugal partition chromatography (CPC) method and applied it to an aqueous C. sativa wood extract. We determined the partition coefficients (KD) of the six major compounds using four butanol-/water-based biphasic solvent systems. Initially, we utilized the n-butanol/propanol/water (3:1:4, v/v/v) systems for the first fractionation step. Subsequently, we employed the water/methyl tert-butyl ether/butanol/acetone (8:5:3:4, v/v/v/v) system to fractionate moderately and highly hydrophilic fractions. We calculated the KD values for major compounds of the most hydrophilic fractions using the butanol/ethanol/water (4:1:5, v/v/v) and butanol/isopropanol/water (2:1:3, v/v/v) systems. In total, we isolated 23 compounds through a combination of CPC, size exclusion chromatography, and preparative HPLC. Among these compounds, six have never been previously described. We characterized them by 1D and 2D NMR experiments and high-resolution mass spectroscopy acquisitions.

Impact of Different Stoppers on the Composition of Red and Rosé Lagrein, Schiava (Vernatsch) and Merlot Wines Stored in Bottle.

Different technical cork stoppers (blend of natural cork microgranules, one-piece natural cork, agglomerated natural cork, technical cork 1+1) were compared to evaluate the effects on the phenolic, volatile profiles and dissolved oxygen in three red (Merlot, Lagrein red, St. Magdalener) and one rosé (Lagrein rosé) wines, which were stored in bottles for 12 months. Gallic acid, caffeic acid, p-coumaric acid, trans-resveratrol, glutahionyl caftaric acid and protocatechuic acid did not vary remarkably during the first three months, whereas at six months a net increase was reported, followed by a clear decrease at 12 months. The same trend was observed in Lagrein rosé, but only for gallic acid. The total anthocyanins content decreased during the storage period in the four wines. Isopentyl acetate, 1-hexanol, ethyl hexanoate, 2-phenylethyl alcohol, diethyl succinate, ethyl octanoate and ethyl decanoate were the main volatile compounds. The sum of alcohols decreased in all four of the wines. The esters decreased in Merlot; however, they increased in Lagrein rosé, Lagrein red and St. Magdalener during the storage. Univariate and multivariate statistics and a sensory discriminant triangle test displayed similar influences of the different stoppers on the phenolic and volatile compounds during the whole storage duration. The changes detected in the phenolic and volatile concentrations were presumably due to the non-oxygen-mediated reactions occurring during 12 months of storage in bottle.

Neuroprotective Effects of Pomegranate Juice against Parkinson's Disease and Presence of Ellagitannins-Derived Metabolite-Urolithin A-In the Brain.

Pomegranate juice is a rich source of ellagitannins (ETs) believed to contribute to a wide range of pomegranate's health benefits. While a lot of experimental studies have been devoted to Alzheimer disease and hypoxic-ischemic brain injury, our knowledge of pomegranate's effects against Parkinson's disease (PD) is very limited. It is suggested that its neuroprotective effects are mediated by ETs-derived metabolites-urolithins. In this study, we examined the capability of pomegranate juice for protection against PD in a rat model of parkinsonism induced by rotenone. To evaluate its efficiency, assessment of postural instability, visualization of neurodegeneration, determination of oxidative damage to lipids and α-synuclein level, as well as markers of antioxidant defense status, inflammation, and apoptosis, were performed in the midbrain. We also check the presence of plausible active pomegranate ETs-derived metabolite, urolithin A, in the plasma and brain. Our results indicated that pomegranate juice treatment provided neuroprotection as evidenced by the postural stability improvement, enhancement of neuronal survival, its protection against oxidative damage and α-synuclein aggregation, the increase in mitochondrial aldehyde dehydrogenase activity, and maintenance of antiapoptotic Bcl-xL protein at the control level. In addition, we have provided evidence for the distribution of urolithin A to the brain.

Crown Procyanidin Tetramer: A Procyanidin with an Unusual Cyclic Skeleton with a Potent Protective Effect against Amyloid-ß-Induced Toxicity.

The structure of a new procyanidin tetramer, which we call a crown procyanidin tetramer, with an unprecedented macrocyclic structure has been characterized for the first time. Its comprehensive spectroscopic analysis revealed that it is a symmetric procyanidin tetramer composed of four (-)-epicatechin sub-units linked alternatively via 4ß→8 or 4ß→6 B-type interflavanyl linkages to form the macrocyclic structure. This NMR-characterized carbon skeleton has never been reported before for procyanidins in grape or in wine, neither in the plant kingdom. Surprisingly, the crown procyanidin tetramer appeared to be specifically localized in grape skin, contrasting with the oligomeric and polymeric procyanidins present in seed, skin, and bunch stem. Moreover, this crown procyanidin tetramer showed promising protective effects against amyloid-ß induced toxicity.

Copigmentation of Malvidin-3-O-Monoglucoside by Oenological Tannins: Incidence on Wine Model Color in Function of Botanical Origin, pH and Ethanol Content.

The effect of the botanical origin, pH level and ethanol content of different oenological tannins on the color of malvidin-3-O-monoglucoside solution, including their effectiveness as copigments, was studied. Briefly, a model wine solution (4 g/L of tartaric acid, pH 3.5 and 12% ethanol) containing 50 mg/L of malvidin-3-O-monoglucoside was prepared and supplemented with 0.1, 0.2 and 0.4 g/L of commercial tannins using (-)-epicatechin as reference control copigment. Six additional model wine solutions (12% ethanol at pH 3.1, 3.5 or 3.9, and 10%, 12% or 14% ethanol at pH 3.5) were prepared as previously described. Samples were stored under airtight conditions. After a week the full absorbance spectrum in the visible range (400-800 nm) was measured and CIELAB color space was determined. These measurements, including an increase in a* (redness), a decrease in b* (yellowness) and a decrease in L* (lightness), indicated that all oenological tannins had a clear positive effect on color copigmentation. Moreover, hydrolysable tannins appeared to be better copigments than condensed tannins as the copigmentation effectiveness (Cp) was found to be between two to four times higher. The effects of these tannins were dose-dependent because a higher addition resulted in a greater impact on copigmentation. In general, an increase in pH and ethanol content resulted in a decrease of the effect of tannins on color. Independent of intrinsic wine conditions, hydrolysable tannins, more specifically gallotannin, remain the most effective in increasing red wine color. These results prove that supplementation with oenological tannins, especially hydrolysable tannins, could be an interesting tool for the improvement of the red wine color.

Comparison of Aquitaine and Rioja Red Wines: Characterization of Their Phenolic Composition and Evolution from 2000 to 2013.

Wine chemical analysis was carried out on 194 commercial blended red wines produced by two major wine-growing areas-the Aquitaine (France) and Rioja (Spain) regions-in order to compare the wines of both regions. Anthocyanins and derived pigments, tannins and derivatives were identified and quantified by HPLC-DAD-ESI-MS/MS (high pressure liquid chromatography coupled to diode array detector and mass spectrometry using the electrospray ionization interface). Mean degree of polymerization (mDP) was determined. The influence of the wine-growing region and the predominance of the properties of some grape varieties used are confirmed by the significant differences observed between both regions. Rioja and Bordeaux "generic" (Bordeaux and Bordeaux-Supérieur appellations) red wines showed the highest anthocyanic content and the highest mDP, as these wines are in a majority made from Merlot (Bordeaux "generic") and Tempranillo (Rioja). On the contrary, Bordeaux "specific" regions (Blayais, Médoc, Graves, and Libournais) showed the red wines with the highest total phenolic content and tannin concentration, as the predominant grape variety used is Cabernet Sauvignon. A principal component analysis (PCA) and a hierarchical ascendant classification (HAC) suggesting patterns between the chemical parameters and the distribution of the red wines in three groups were proposed. The comparison of the two wine-growing areas also reveals some similarities between the various grape varieties used. A general effect of a progressive decrease in anthocyanins, anthocyanin-derived pigment and tannins is observed for older wines.

In Vitro Glucuronidation and Sulfation of ε-Viniferin, a Resveratrol Dimer, in Humans and Rats.

ε-Viniferin is a resveratrol dimer that possesses antioxidant or anti-inflammatory activities. However little is known about the metabolism of this oligostilbene. This study was thus undertaken as a first approach to identify and characterize the metabolites of ε-viniferin and to describe the kinetic profile of their appearance in humans and rats. The glucuronides and sulfates of ε-viniferin were first obtained by chemical hemi-synthesis and were fully characterized by UPLC-MS and NMR spectroscopy. Then, ε-viniferin was incubated with human or rat S9 liver fractions that led to the formation of four glucuronoconjugates and four sulfoconjugates. In both species, ε-viniferin was subjected to an intense metabolism as 70 to 80% of the molecule was converted to glucuronides and sulfates. In humans, the hepatic clearance of ε-viniferin (Vmax/Km) for glucuronidation and sulfation were 4.98 and 6.35 µL/min/mg protein, respectively, whereas, in rats, the hepatic clearance for glucuronidation was 20.08 vs. 2.59 µL/min/mg protein for sulfation. In humans, three major metabolites were observed: two glucuronides and one sulfate. By contrast, only one major glucuronide was observed in rats. This strong hepatic clearance of ε-viniferin in human and rat could explain its poor bioavailability and could help to characterize its active metabolites.

Structures of polymeric pigments in red wine and their derived quantification markers revealed by high-resolution quadrupole time-of-flight mass spectrometry.

RATIONALE: The quantification of polymeric pigment families in red wine through their derived quantification markers released during phloroglucinolysis will allow the understanding of their formation kinetics and evolutions during aging which has not been achieved until now and is in urgent need. The identification of these quantification markers was achieved by high-resolution mass spectrometry (HRMS) in this study. METHODS: HRMS was used to clarify the fragmentation patterns in positive mode of polymeric pigments and identify their derived quantification markers released during phloroglucinolysis. RESULTS: With HRMS, identification of (epi)catechin-malvidin-3-O-glucoside adducts was simplified to MS/MS, and the fragmentation pattern of malvidin-3-O-glucoside-(epi)catechin adducts was clearly demonstrated. The attribution of four detected ions at m/z 1071.2765 in red wine to the trimeric structure of (epi)catechin-[malvidin-3-O-glucoside-A type linkage-(epi)catechin] and [malvidin-3-O-glucoside-A type linkage-(epi)catechin]-(epi)catechin was achieved for the first time by MS/MS and evidence given by phloroglucinolysis. Moreover, four kinds of derived quantification markers released from polymeric pigments during phloroglucinolysis were also identified. CONCLUSIONS: The fragmentation pathways of polymeric pigments in red wine and their derived quantification markers released during acidic chemical depolymerisation were clarified which will allow their quantification in red wine.

Hypotheses on the effects of enological tannins and total red wine phenolic compounds on Oenococcus oeni.

Lot of articles report on the impact of polyphenols on wine lactic acid bacteria, but it is clear that the results still remain confusing, because the system is complicated both in term of chemical composition and of diversity of strains. In addition, red wines polyphenols are multiple, complex and reactive molecules. Moreover, the final composition of wine varies according to grape variety and to extraction during winemaking. Therefore it is nearly impossible to deduce their effects on bacteria from experiments in oversimplified conditions. In the present work, effect of tannins preparations, currently considered as possible technological adjuvants, was assessed on growth and malolactic fermentation for two malolactic starters. Experiments were conducted in a laboratory medium and in a white wine. Likewise, impact of total polyphenolic extracts obtained from different grape variety red wines was evaluated in the white wine as culture medium. As expected growth and activity of both strains were affected whatever the additions. Results suggest some interpretations to the observed impacts on bacterial populations. Influence of tannins should be, at least partly, due to redox potential change. Results on wine extracts show the need for investigating the bacterial metabolism of some galloylated molecules. Indeed, they should play on bacterial physiology and probably affect the sensory qualities of wines.

Chip electrophoresis as a novel approach to measure the polyphenols reactivity toward human saliva.

Saliva is a biological fluid with a multifunctional role that makes it interesting in terms of research and diagnostic possibilities. In food research, human saliva represented a useful tool by which we measure the tactile sensation elicited by polyphenol-rich beverages called astringency. A method based on SDS-PAGE analysis of saliva before and after the binding reaction with wine polyphenols has been successfully used in previous studies for measuring wine astringency by means of the saliva precipitation index. In this work, chip electrophoresis was used alternatively to SDS-PAGE and results were compared. Chip electrophoresis provides a very good reproducibility for wine and grape astringency. Moreover, this approach is much faster than the conventional SDS-PAGE method requiring several hours for an analysis. Another advantage over traditional gel is lower sample and reagent volume requirements, as well as the lower and less toxic wastes, contributing benefits to health and environment. The application of this novel method allowed, using the principal component analysis, to distinguish grapes and wines according to the saliva precipitation index and structural characteristics determined by the phoroglucinolysis analysis.

Soil effect on proanthocyanidins composition of red and white wines obtained from Nero d'Avola and Grillo Vitis vinifera L. Cultivars.

In this study, the effects of the main soil chemical-physical parameters (i.e. texture, pH, total carbonates, cation exchange capacity, electric conductivity, organic matter and mineral endowment) on proanthocyanidin composition of Nero d'Avola red wines and Grillo white wines were investigated. Monomer proanthocyanidins (i.e. (+)-catechin and (-)-epicatechin) and oligomer proanthocyanidins (i.e. B1, B2, B3 and B4 dimers and C1 trimer), as well as proanthocyanidins subunit composition, percentage of galloylation, percentage of prodelphinidins and mean degree of polymerization, were studied for each wine. Results highlighted that the proanthocyanidins composition of both red and white wines is greatly affected by soil. In particular, the proanthocyanidins composition of Nero d'Avola red wines appeared to be affected by the soil physical-chemical parameters related to nutrients dynamics (CEC, EC, pH, organic matter, mineral endowment), whereas the proanthocyanidins composition of Grillo white wines was mainly influenced by the soil texture, that modulates soil water dynamics.

Comparative evaluation of the phenolic content and antioxidant capacity of sun-dried raisins.

BACKGROUND: Raisins are one of the favorite dried fruit because of their high healthful and nutrimental values. Three white (Besni beyazi-BBR, Hatun parmagi-HPR and Sultaniye-SR) and two red (Antep karasi-AKR and Besni karasi-BKR) grape varieties were used in the present study. The aim of this study was to determine and compare the phenolic composition and antioxidant properties of Turkish raisins. RESULTS: Four flavan-3-ols, six phenolic acids, four flavonols and 13 anthocyanins were identified and quantified in raisins. (+)-Catechin (range, 56.3-419 mg kg⁻¹) was the mosst abundant flavanol, trans-caftaric acid (range, 20.48-114 mg kg⁻¹) was the abundant dominannt phenol acid, quercetin-3-O-glucoside (range, 2.79-12.83 mg kg⁻¹) was the dominant flavonoll and malvidin-3-O-(6-O-p-coumaroyl)-glucoside (range, 16.75-22.59 mg kg⁻¹) wass the major anthocyanin in all raisins. Antioxidant capacity were 22.69-63.66, 5.07-40.47, 7.00-17.69 and 40.d 40.74-77.41 mmol Trolox kg⁻¹ as determined bby the ABTS, DPPH, FRAP and ORAC assays, respectively. CONCLUSIONS: 27 phenolic compounds from four phenolic families (i.e. flavan-3-ols, phenolic acids, flavonols and anthocyanins) have been characterised in the raisins of three white and two red grapes. The total phenolic and anthocyanins content vary widely among different raisins. Strong correlations between antioxidative capacity and phenolic content and between antioxidative capacity and flavonol content were noticed.

Color Characterization of Bordeaux Red Wines Produced without Added Sulfites.

Nowadays, the development of naturalness as a concept is illustrated in the oenological field by the development of wine produced with lower inputs, sometimes even without any addition of SO2 throughout the winemaking process, up to the bottling stage. Despite the increase in the offer of these wines, they remain poorly explored in the literature and require characterization. This study was developed to evaluate the color of Bordeaux red wines without SO2 addition using colorimetric and polymeric pigments analysis. From a batch of commercial Bordeaux red wines with and without SO2 addition, and experimental wines produced from homogenous grapes according to different winemaking processes, colorimetric analyses (CIELab and color intensity (CI)) revealed a large difference in wine color depending on the presence or absence of SO2. Indeed, wines without SO2 were significantly darker and presented with a deeper purplish color. According to these observations, polymeric pigments were quantified using UPLC-DAD/ESI QTof, and a higher concentration of polymeric pigments bound by the ethylidene bridge was observed in wines without SO2. This correlated with differences observed for CIELab and CI. Finally, a comparison with polymeric tannins bound by ethylidene bridge was made and revealed that no differences were observed between wines with and without added SO2. This underlines the affinity difference between tannins and anthocyanins to react with acetaldehyde to form ethylidene bridges.

Evolution of the Crown Procyanidins' Tetramer during Winemaking and Aging of Red Wine.

Condensed tannins play a major role in the quality of red wine. After grape extraction, they quickly evolve thanks to different oxidation mechanisms. Recently, NMR identified a new sub class of condensed tannins, named crown procyanidins, in red wine. The crown procyanidins' tetramer exhibits a macrocyclic structure composed of four (-)-epicatechin with an unusual cavity in the center of the molecule. These new tannins exposed a higher polarity than the linear tannins. In this work, the evolution kinetics of these crown procyanidins during the winemaking process and after aging of red wine in bottles were studied. Samples' quantification was analyzed by UPLC-UV-Q-TOF. The concentration of cyclic and non-cyclic procyanidins was compared. During the winemaking process, crown procyanidins are mainly extracted at the beginning of the alcoholic fermentation and they remain stable until the end of the winemaking process. The high polarity and solubility of this new molecule in water was confirmed. During the aging of red wine in bottles, crown procyanidins' concentrations are stable, whereas the non-cyclic tannins decrease dramatically. Finally, a strong oxygenation experiment confirmed the crown procyanidins' resistance to oxidation and unique skills.

Identification, quantitation and sensory contribution of new C-glucosidic ellagitannin-derived spirit compounds.

Ellagitannins are the main extractible phenolic compounds in oak wood. They can be extracted by eaux-de-vie aging in barrels but rapidly undergo chemical transformations, such as hydrolysis, reduction, oxidation, or even substitution reactions. Given gaps in knowledge about their composition in spirits, the goal of this work was to explore ellagitannin behavior during Cognac eaux-de-vie aging and search for new C-glucosidic ellagitannin-derived compounds. A purification protocol was followed to isolate the new formed compounds. UPLC-UV-Q-ToF and NMR spectroscopy were used to characterize a new C-glucosidic ellagitannin, named brandy tannin B. Two new forms of whiskey tannins were also isolated. The new compound, Brandy tannin B along with ß-1-O-ethylvescalagin and ß-1-O-ethylvescalin, and whiskey tannins were identified and quantified for the first time in Cognac eaux-de-vie. In addition, the organoleptic impact of brandy tannin B was evaluated. This work brings new insights into ellagitannin composition and their contribution to spirit quality.

Oxyresveratrol and Gnetol Glucuronide Metabolites: Chemical Production, Structural Identification, Metabolism by Human and Rat Liver Fractions, and In Vitro Anti-inflammatory Properties.

Stilbene metabolites are attracting great interest because many of them exhibit similar or even stronger biological effects than their parent compounds. Furthermore, the metabolized forms are predominant in biological fluids; therefore, their study is highly relevant. After hemisynthesis production, isolation, and structural elucidation, three glucuronide metabolites for oxyresveratrol (ORV) were formed: trans-ORV-4'-O-glucuronide, trans-ORV-3-O-glucuronide, and trans-ORV-2'-O-glucuronide. In addition, two glucuronide metabolites were obtained for gnetol (GN): trans-GN-2'-O-glucuronide and trans-GN-3-O-glucuronide. When the metabolism of ORV and GN is studied in vitro by human and rat hepatic enzymes, four of the five hemisynthesized compounds were identified and quantified. Human enzymes glucuronidated preferably at the C-2' position, whereas rat enzymes do so at the C-3 position. In view of these kinetic findings, rat enzymes have a stronger metabolic capacity than human enzymes. Finally, ORV, GN, and their glucuronide metabolites (mainly at the C-3 position) decreased nitric oxide, reactive oxygen species, interleukin 1ß, and tumor necrosis factor α production in lipopolysaccharide-stimulated macrophages.

Antisense down-regulation of 4CL expression alters lignification, tree growth, and saccharification potential of field-grown poplar.

Transgenic down-regulation of the Pt4CL1 gene family encoding 4-coumarate:coenzyme A ligase (4CL) has been reported as a means for reducing lignin content in cell walls and increasing overall growth rates, thereby improving feedstock quality for paper and bioethanol production. Using hybrid poplar (Populus tremula × Populus alba), we applied this strategy and examined field-grown transformants for both effects on wood biochemistry and tree productivity. The reductions in lignin contents obtained correlated well with 4CL RNA expression, with a sharp decrease in lignin amount being observed for RNA expression below approximately 50% of the nontransgenic control. Relatively small lignin reductions of approximately 10% were associated with reduced productivity, decreased wood syringyl/guaiacyl lignin monomer ratios, and a small increase in the level of incorporation of H-monomers (p-hydroxyphenyl) into cell walls. Transgenic events with less than approximately 50% 4CL RNA expression were characterized by patches of reddish-brown discolored wood that had approximately twice the extractive content of controls (largely complex polyphenolics). There was no evidence that substantially reduced lignin contents increased growth rates or saccharification potential. Our results suggest that the capacity for lignin reduction is limited; below a threshold, large changes in wood chemistry and plant metabolism were observed that adversely affected productivity and potential ethanol yield. They also underline the importance of field studies to obtain physiologically meaningful results and to support technology development with transgenic trees.

Identification, amounts, and kinetics of extraction of C-glucosidic ellagitannins during wine aging in oak barrels or in stainless steel tanks with oak chips.

The C-glucosidic ellagitannins are found in wine as a result of its aging in oak barrels or in stainless steel tanks with oak chips. Once dissolved in this slightly acidic solution, the C-glucosidic ellagitannins vescalagin can react with nucleophilic entities present in red wine, such as ethanol, catechin, and epicatechin, to generate condensed hybrid products such as the ß-1-O-ethylvescalagin and the flavano-ellagitannins (acutissimin A/B and epiacutissimin A/B), respectively. During this study, we first monitored the extraction kinetic and the evolution of the eight major oak-derived C-glucosidic ellagitannins in red wines aged in oak barrels or in stainless steel tank with oak chips. Their extraction rates appeared to be faster during red wine aging in stainless steel tanks with oak chips. However, their overall concentrations in wines were found higher in the wine aged in barrels. The formation rates of the vescalagin-coupled derivatives were also estimated for the first time under both red wine aging conditions (i.e., oak barrels or stainless steel tanks with oak chips). As observed for the oak-native C-glucosidic ellagitannins, the concentrations of these vescalagin derivatives were higher in the red wine aged in oak barrels than in stainless steel tanks with oak chips. Despite these differences, their relative composition was similar under both red wine aging conditions. Finally, the impact of the oak chips size and toasting level on the C-glucosidic ellagitannins concentration in wine was also investigated.

Chemical and sensory evaluation of Bordeaux wines (Cabernet-Sauvignon and Merlot) and correlation with wine age.

Wine quality study was carried out with 24 vintages of Cabernet-Sauvignon (CS) and 7 vintages of Merlot (M) produced by two different Bordeaux wine-growing areas. Proanthocyanidin monomers and oligomers were identified and quantified by HPLC-UV-Fluo. Percentage of galloylation (%G), of prodelphinidins (%P) as well as mean degree of polymerisation (mDP) were also determined. Total phenolic compounds, total anthocyanins, total tannins, hue, CI (colour intensity), titratable acidity, ethanol level and pH were evaluated. Sensory analysis concerning astringency and bitterness intensity was also performed. Total phenolic compounds, total anthocyanins, total tannins, tannin monomers, hue, CI, % G, % P, mDP and astringency intensity differentiate both wines (M and CS) according to vintage. Correlations between wine age and: mDP, hue, astringency and tannin monomers (C+EC) are obtained. Qualitative tannin characterisation is established by correlation between astringency and mDP (R(2)=0.509, p=0.051, CS; R(2)=0.780, p=0.000M). In addition, mDP decreases significantly during ageing (R(2)=0.796, p=0.000; CS and R(2)=0.946, p=0.000; M). Scale patterns between wine mDP and tannin perception (astringency) are proposed.

Effect of enological treatments on phenolic and sensory characteristics of red wine during aging: Micro-oxygenation, sulfur dioxide, iron with copper and gelatin fining.

This work aimed to study long-term impact of micro-oxygenation and/or different aging treatments: (i) high SO2, (ii) high Fe with Cu and (iii) gelatin fining on Plavac mali red wine phenolic and in-mouthfeel sensory development in barrels and furthermore in bottles. Results showed that outcomes of micro-oxygenation strongly depend on aging treatments. High SO2 concentration during aging in barrels and bottles delayed typical phenolic changes and slightly contributed to astringency and lower color intensity, particularly in wine that was not micro-oxygenated. High metal concentrations and gelatin fining promoted intensive polymerization of proanthocyanins and a lower percentage of prodelphinidins after long-term aging in barrels. Also, flavan-3-ol and anthocyanins transformation rates in micro-oxygenated wines of both treatments significantly differed from their controls. Gelatin fining proved to be a very effective treatment for astringency reduction, particularly when combined with micro-oxygenation, but fined wines after long term aging in bottles showed lower color intensity.