Maximizing Wine Antioxidants: Yeast’s Contribution to Melatonin Formation

Melatonin is commonly found in various fruits, juices, and some fermented beverages. Its concentration in wine is influenced by soil properties, climatic factors, and yeast activity. Even if it is found in fermented beverages in relatively low proportions, melatonin still holds significant nutritional value, giving anti-aging properties, anti-inflammatory actions, and antidepressant effects. In this context, this article focuses on evaluating the impact of different Saccharomyces and non-Saccharomyces yeast species on the formation of melatonin and its contribution to wines’ total antioxidant capacity. Considering that the antioxidant activity of wine is usually related to the content of phenolic compounds, ten such compounds were analyzed. The evaluation of bioactive compounds was performed using high-performance liquid chromatography (HPLC) coupled with mass spectrometry. The total antioxidant capacity of wine samples was evaluated by the ABTS+ method. The administration of bâtonnage products increased the efficiency of non-Saccharomyces yeasts. The mixtures of Saccharomyces and non-Saccharomyces yeasts generated higher values for melatonin. The results confirm a significant impact from the grape variety and the specific yeast strains on the melatonin concentration. Also, a strong dependence between antioxidant activity and melatonin levels was observed. Given the limited existing studies on the presence of melatonin in wines, new perspectives are needed for future exploration and understanding.


Introduction
Melatonin, which is typically synthesized from tryptophan, has received great attention in recent years due to its potential antioxidant activity.In plants, melatonin promotes growth and has shown anti-senescence effects, while in the human body, it helps to improve circadian rhythms, sustain eye health and neurological activity, and has manifested antiinflammatory, anti-cancer, anti-diabetes, and anti-aging properties.Moreover, melatonin contributes to a general state of well-being, reducing anxiety and depression [1].The ability to synthesize melatonin in the human body diminishes with age and is influenced by the adopted lifestyle.Thus, diet is an essential factor in the synthesis of different bioactive compounds, including melatonin [2].This compound can be found in a variety of seeds, cereals, and fruits, including strawberries, blueberries, cherries, sour cherries, and table grapes.It was also identified in drinks, including fruit juices, coffee, tea, beer, and wine [3][4][5][6].However, relatively few studies have focused on the melatonin content of wine.Although the nutraceutical effects of wine consumption are a controversial subject due to the alcohol content, wine is made up of numerous classes of compounds, many of which have positive effects on the human body.Therefore, the tendency is to produce wines with an improved nutritional value, to counterbalance the negative impact of alcohol.In this context, Boban et al. [7] demonstrated the protective action of wine on the cardiovascular system.In this sense, following an induced myocardial infarction, the survival rate was significantly higher in rats that received reduced amounts of white wine (72.2%), compared to those that received water (47.8%).This effect can be attributed to the numerous biologically active compounds, especially phenolic compounds and melatonin [8,9].
The majority of the existing studies correlate the antiradical activity in wines with their phenolic compound profile [10].Some research articles that focused on the impact of moderate consumption (1-2 glasses/day) of wine reported a significant increase in the antioxidant capacity in plasma, high-density lipoprotein levels, in parallel with an important decrease in oxidative stress, cardiovascular diseases and cancer cells [11,12].Different Saccharomyces spp.yeasts (Saccharomyces cerevisiae, Saccharomyces uvarum) and non-Saccharomyces spp.(Candida colliculosa, Candida stellata, Metschnikowia pulcherima, Torulospora delbrueckii, Kloeckera thermotolerans, Kloeckera apiculata, Hanseniaspora uvarum) were isolated in wine fermentation [13].In general, non-Saccharomyces types of yeast cannot complete alcoholic fermentation and so are frequently used in combination with Saccharomyces yeasts.Non-Saccharomyces species are able to reduce the initial ethanol level by 1-2% v/v.Gomez et al. [14] studied the evolution of melatonin and its isomer in the Malbec grape variety using an UHPLC-MS/MS system.Melatonin was identified in the grape extract while its isomer was present in musts and wines.The results confirmed that Saccharomyces cerevisiae plays an essential role in the production of melatonin and its isomer in wine.Fernández-Cruz et al. [15] studied melatonin and derived tryptophan metabolites produced during alcoholic fermentation by different Saccharomyces and non-Saccharomyces (Torulaspora delbrueckii and Metschnikowia pulcherrima) yeast strains.In Romania, Albu et al. [16] reported for the first time the analysis of melatonin and its precursors.Their study focuses on the development and validation of a sensitive and selective HPLC-MS/MS method for the simultaneous analysis of melatonin, serotonin, and tryptophan in wine samples.Rodriguez-Naranjo et al. [17] did not identify melatonin in Cabernet Sauvignon, Merlot, Syrah, Tempranillo, Tintilla de Rota, Petit Verdot, Pedro Ximénez, Nebbiolo, Palomino Fino.Also, the same results were presented for Flame Seedless, Red Globe, Moscatel Italica, and Superior Seedless, all table grapes from Spain.In a previous study published by the authors [18], melatonin was found in important amounts in table grapes from Romania (Timpuriu de Pietroasa, Coarnă neagră select , ionată, Paula).In general, dosages of 0.5 to 5 mg are well tolerated and have no side effects.Although melatonin is naturally present in plants, the quantities are extremely low, making it impossible to obtain concentrations that exceed the maximum permissible concentration, which is the limit for harmful effects on the human body.[19].Indeed, the concentration of melatonin in fermented beverages is usually low (pg/mL to ng/L), but manifests an important contribution to their nutritional value, which has been less studied in white wines [20].The increase in the consumption of white wine at the global level requires the development of additional research on the biological effects of white wine [7].There is increased interest in the finding of natural sources of melatonin and studies are not sufficient in this area.The goal of the present research is to obtain wines that support balanced diets, with high antioxidant capacity.For this reason, this study focuses on optimizing the production technology of some wines by monitoring the influence of different yeasts (Saccharomyces and non-Saccharomyces yeasts) on the production of melatonin.Since most authors report the antioxidant activity of wines according to the presence of phenolic compounds, ten such compounds were analyzed.To amplify the yeasts' activity, some bâtonnage products were also applied.The topic is up-to-date and presents novelty through the chosen varieties, but also through the diversity of the inoculated yeasts and applied technology.

Phenolic Compound Evaluation
The principal phenolic compounds were determined using an Agilent 1100 HPLC Series system (Agilent, Santa Clara, CA, USA) coupled with an Agilent Ion Trap VL mass spectrometer (Agilent, Santa Clara, USA), following the method presented in our team's previous papers [21,22].Wine samples were filtered using 0.45 µm sterile filters.Determinations were performed in triplicate and the results are presented as means, including between 0.3 g/L (S12) and 1.9 g/L (S27), while the alcoholic concentration was between 9.5% (S14) and 12.3% (S18, S21) alc.vol.

Phenolic Compound Evaluation
The principal phenolic compounds were determined using an Agilent 1100 HPLC Series system (Agilent, Santa Clara, CA, USA) coupled with an Agilent Ion Trap VL mass spectrometer (Agilent, Santa Clara, USA), following the method presented in our team's previous papers [21,22].Wine samples were filtered using 0.45 µm sterile filters.Determinations were performed in triplicate and the results are presented as means, including

Phenolic Compound Evaluation
The principal phenolic compounds were determined using an Agilent 1100 HPLC Series system (Agilent, Santa Clara, CA, USA) coupled with an Agilent Ion Trap VL mass spectrometer (Agilent, Santa Clara, USA), following the method presented in our team's previous papers [21,22].Wine samples were filtered using 0.45 µm sterile filters.Determinations were performed in triplicate and the results are presented as means, including standard deviations.For the analyzed phenolic compounds, the detection limit (LOD) was 0.04 µg/mL, while the quantification limit (LOQ) was 0.2 µg/mL.Some analytical conditions are presented in Table 2.This analysis was performed according to the method presented in the previous paper [18], with some modifications and using a Transcend XT Ultimate 3000 UHPLC system (Thermo Scientific TM, Waltham, MA, USA) coupled with a TSQ Access Max mass spectrometer.The elution of compounds was carried out using an Agilent Poroshell C18 column (Agilent, USA) (4.6 × 100 mm, 1.8 µm).The transfer was achieved using a 30% water (TA) and 70% methanol (TD) mixture (v/v) (in loop).The TX column was prepared using a mixture (v/v/v) of acetonitrile (45%), isopropanol (45%), and acetone (10%) (TB), followed by triethylamine (0.05%) solution prepared in acetonitrile (TC).For separating melatonin from the internal standard, a mobile phase of 0.1% formic acid in water (LA) and 0.1% formic acid in methanol (LB) was used.Regarding the mass spectrometry analysis, a heated electrospray ionization source (in positive mode) was used for ionization, while a collision cell within Q2 was engaged in fragmenting and separating particular ions to allow accurate identification.The ionization conditions were set with an ionization potential of 3 kV, an ionization source temperature of 350 • C, a nebulization gas pressure of 35 psi, and an auxiliary gas pressure of 10 psi.The capillary tube was kept at 350 • C, while the polarity was positive [18].
For the standard solutions, a quantity of 10 mg of melatonin was dissolved in methanol and diluted to 10 mL in the same solvent.A volume of 0.5 mL solution was diluted to 10 mL with methanol.A volume of 0.1 mL from the previous solution was diluted with 10 mL of methanol.An internal standard was prepared in water by dissolving a quantity of 6 mg of tryptophan in 10 mL of solvent.A volume of 1.66 mL was diluted to 5 mL with methanol.A series of concentrations of 0.5, 2.0, 10.0, 25.0, and 100 ppb were produced by taking corresponding volumes of the stock solutions, a 0.05 mL internal standard, and dilution to 1 mL with a mixture of water: methanol (50%:50% (v/v)).For sample preparation, a volume of 2.5 mL of wine was filtered through a 0.45 µm nylon filter; a portion of 1 mL of clear filtrate was spiked with 0.05 mL of internal standard and subjected to analysis according to the method.For the control samples, the second volume of 1.0 mL of filtrate was subjected to analysis to subtract the tryptophan content from the sample.The areas of tryptophan were subtracted from the control samples and used further for the calculation of melatonin in the wine samples.The method's selectivity was confirmed using blank solutions, showing no interference.Melatonin detection was based on the specific transition of m/z 233−→174, with a retention time at 3.28 min.For the internal standard, the detection used the transition of m/z 205.1−→146, with a retention time at 2.50 min.A supplementary chromatographic peak was observed in the melatonin chromatogram, likely due to a tryptophan impurity, but it did not interfere with melatonin determination.The method demonstrated linearity in the range of 0.05 ppb to 100 ppb, with calibration points at 0.5, 2.0, 10.0, 25.0, and 100 ppb.The regression correlation coefficient was 0.9945.The back-calculation of standard concentrations using the regression equation showed values within 85% to 115% of the expected concentrations.The highest deviation was 5.85% at 0.5 ppb, and the lowest was 0.7% at 100 ppb.The standard relative deviation for three series of samples under the same conditions was 1.5%.The LOD and LOQ were calculated using the standard deviation of the intercept and the slope, multiplied by 3.3 for LOD and 10 for LOQ.The LOQ was 0.12 ppb, and the LOD was 0.059 ppb, confirmed by the signalto-noise ratio.Accuracy and precision were evaluated using standard method addition at concentrations of 1 ppb, 25 ppb, and 100 ppb in representative wine samples.Recovery rates were within 85% to 115%: 87.3% at 1 ppb, 92% at 25 ppb, and 93.2% at 100 ppb.Repeatability was assessed with three concentrations within the linearity range, achieving values within 98% to 102% of the target.Inter-day and intra-day precision showed values of 8.5% for the lowest concentration and 6.5% for the highest concentration.The samples were analyzed in triplicate and the results are presented as arithmetic means and standard deviations.The concentration of melatonin is expressed in µg/L [18].

Total Antioxidant Capacity
Total antioxidant capacity of wine samples was evaluated by ABTS + method (also known as Trolox equivalent antioxidant capacity (TEAC) assay), which relies on the ability of antioxidants to diminish the blue-green color of ABTS + in correspondence with their concentrations and scavenging properties.Initially, the reduced ABTS molecule is converted by oxidation to ABTS + using hydrogen peroxide-H 2 O 2 in an acidic medium of 30 mmol/L acetate buffer solution (pH = 3.6).In the acetate buffer solution, the concentrate (deep green) ABTS + molecules persist for a long time.Another solution of 0.4 mmol/L acetate buffer (pH = 5.8) was prepared and used for the dilution of the initial medium.The color of ABTS + molecules was spontaneously and gradually decolorized.The decolorizing rate is proportional to the concentrations in different antioxidant compounds.The absorbance was monitored at 660 nm and the antioxidant capacity is inversely related to the decolorizing rate of the mixture.The calibration curve was made with Trolox solution and the results are expressed as mmol Trolox equivalent per liter [23].

Statistical Tests
The statistical analysis of the data was carried out using XLStat (Luminevo, Denver, CO, USA) and aimed at the analysis of variance (ANOVA) which reveals the existence of a statistically significant difference (p-value < 0.05) between the analyzed samples.Student-t test highlights pairs of samples that are significantly different from each other (p-value < 0.05).The possible existing correlations between the analyzed bioactive compounds were highlighted by principal components analysis (PCA).Linear regression analysis highlighted the influence of the analyzed bioactive compounds on the antioxidant capacity value.

Effect of Different Yeasts on Wine Bioactive Compounds
According to Tables 3 and 4, the samples showed different values of bioactive compounds in relation to the specificity of the grape varieties and the applied technology (different species of inoculated yeasts and various bâtonnage products).Tables S1-S3 contains the differences between each pair of samples, for each bioactive compound.In general, caftaric acid is the main representative in samples obtained from the mix of Aligote + Fetească albă grapes, without bâtonnage (from 16.14 ± 0.15 µg/L in samples with Lachancea thermotolerans yeast-S2 to 19.90 ± 0.15 µg/L in S3-Saccharomyces cerevisiae).This compound is caffeic acid's ethyl ester.The results are in accordance to Peréz-Navarro [24] that presented caftaric acid as one of the predominant phenolic acids in white wines.Its concentrations decreased by up to eight times in the case of samples with bâtonnage (from 2.49 ± 0.01 µg/L in S10-Saccharomyces cerevisiae yeast + bâtonnage products to 3.28 ± 0.00 µg/L in S11-Torulaspora delbrueckii yeast + bâtonnage products).Indeed, bâtonnage (inactivated Saccharomyces cerevisiae yeasts, glutathione and pectolytic enzymes) can increase the wines' complexity and mouthfeel by favoring yeast autolysis and releasing aroma constituents.Other factors that can cause the reduction in phenolic compounds are different chemical and physical processes that can occur, including oxidation, binding to lees, polymerization and precipitation.Certain phenolic molecules have the potential to react with sulfur dioxide, creating more stable complexes [25].Following bâtonnage application to this category of samples, caffeic acid became predominant in most samples (from 9.31 ± 0.12 µg/L in S11-Torulaspora delbrueckii yeast to 10.32 ± 0.01 µg/L in S13-Saccharomyces cerevisiae + Kluyveromyces thermotolerans yeasts + bâtonnage products).This compound usually derives from p-coumaric acid (which results from cinnamic acid), but free forms of caffeic acid can arise due to esterase activity, too [24].Contrary to these results, this compound was not identified in the white wine samples studied by Onache et al. [26], while a strong positive correlation of caffeic acid with catechin, epicatechin and transresveratrol was shown by the authors.In the present article, only the positive correlation between epicatechin and catechin was confirmed by principal component analysis, for both Aligoté + Fetească regală (r = 0.937) and Sauvignon blanc wines (r = 0.762).
Gallic acid showed the highest values in samples obtained from the Sauvignon blanc variety.Garrido and Borges [27] suggested that gallic acid was a significant phenolic compound due to its important scavenging activity.While it can originate from the grape, its presence may also stem from chemical transformations occurring during fermentation.Thus, enzymes and acids present in the grape and microbial activity may catalyze the hydrolysis of hydrolysable and condensed tannins, leading to the release of gallic acid [28].In this category, samples subjected to bâtonnage (from 20.65 ± 0.03 in S27-Saccharomyces cerevisiae + Kluyveromyces thermotolerans yeasts + bâtonnage products to 22.96 ± 0.08 in S23-Lachancea thermotolerans yeast + bâtonnage products) with various oenological products indicating slightly higher concentrations compared to those without bâtonnage (from 18.85 ± 0.02 in S16-Lachancea thermotolerans yeast, to 20.70 ± 0.20 in S21-Kluyveromyces thermotolerans + Torulaspora delbrueckii + Saccharomyces cerevisiae yeasts).If gallic, caffeic, and caftaric acids are predominant in samples without bâtonnage treatment, samples with bâtonnage showed the highest values of gallic acid, caffeic acid, and p-coumaric acid.In a separate study conducted by our team [21], it was observed that enzyme preparations had a notable impact on the generation of various phenolic compounds in Sauvignon blanc wines.Among these compounds, protocatechuic acid and caftaric acid were found to be most predominant.
The analyzed phenolic acids emerged as significant indicators for distinguishing the analyzed varieties across the diverse wine-growing regions in Romania [29].Lengyel [30] also noted comparable concentrations of phenolic compounds in wines derived from Sauvignon blanc varieties.
The content of the samples in bioactive compounds was also evaluated after the application of the bâtonnage products, and Table S3 highlights the differences between the pairs of samples with and without this treatment.Therefore, for Aligoté + Fetească albă wines, the most differences were between the variants S1-S8 (control samples) and S2-S9 (Lachancea thermotolerans vs. Lachancea thermotolerans yeasts + bâtonnage products).Sauvignon blanc samples displayed most differences between the S18-S25 (Torulaspora delbrueckii) and S19-S26 (Pichia kluyveri) pairs.
Certain strains of yeast have the ability to synthesize melatonin from tryptophan during fermentation, although other microorganisms such as bacteria and fungi may also contribute to its synthesis through enzymatic processes [31,32].From Table 3 and Table S1, it can certainly be confirmed that the applied technology (different yeasts, application of bâtonnage) influences the melatonin concentration in Aligote + Fetească albă wines.Yeasts administered in samples S2 (Lachancea thermotolerans yeast) and S7 (Kluyveromyces thermotolerans + Torulaspora delbrueckii + Saccharomyces cerevisiae) did not show significant differences compared to the control sample.With the application of the bâtonnage products, the S14 variant (Kluyveromyces thermotolerans + Torulaspora delbrueckii + Saccharomyces cerevisiae yeasts + bâtonnage products) presented a significant difference from S8 (control sample, with bâtonnage, no exogenous yeasts).
According to Fernández-Cruz et al. [15], each yeast type has the ability to produce melatonin at different growth stages.Alcohol content can influence the dilution and release of phenolic compounds and melatonin [31].For the analyzed samples, minor differences in alcoholic strength were registered.The correlation between melatonin production and growth phase suggests that melatonin may play a role in the yeast's adaptability to the changing conditions of alcoholic fermentation.Also, melatonin-protein binding for some yeast species should be taken into consideration (not analyzed in this paper) [32].Fracassetti et al. [33] identified between 0.038 µg/L and 0.063 µg/L melatonin in red wines, being in accordance with the results presented by Vitalini et al. [34].This compound was found in great amounts (0.011-0.019 µg/mL) in Riesling wines from Romania (commercial samples), analyzed by Albu et al. [16].In another study, Eremia et al. [8] reported 0.74-0.84ng/mL melatonin in Fetească neagră and Cabernet sauvignon red wine samples, comparable with the team's results for white samples.It is clear that different yeasts can synthetize different amounts of bioactive compounds [17].In accordance with Sunyer-Figueres et al. [35], melatonin acts as a modulator of the biosynthesis of different phenolic compounds.In correlation with Morcillo-Parra et al. [32], melatonin increases the survival of non-Saccharomyces species when fermentation is carried out using a mixed inoculum, which is either solely or co-inoculated with non-Saccharomyces and Saccharomyces.Valera et al. [36] suggested that yeast cells become more fermentative in the presence of melatonin, completing the fermentation a day or two sooner.The authors observed that when melatonin was added to the synthetic must, Torulaspora delbrueckii and Saccharomyces bacillaris remained until the completion of the fermentation, but Metschnikowia pulcherrima and Hanseniaspora uvarum only showed up at the start of the process.Therefore, variations in melatonin-protein interactions between non-Saccharomyces species may be explained by variations in sugar metabolism and enzyme activity.In another study, Rodriguez-Naranjo et al. [17] evaluated the ability of several Saccharomyces and non-Saccharomyces yeasts to produce melatonin.Different strains exhibited varying degrees of production; the non-Saccharomyces yeast with the greatest concentration was Starmerella bacillaris.However, depending on the yeast strain, extracellular melatonin was found at various stages of the fermentation process.Nevertheless, the same authors also postulated that melatonin requires tryptophan to be present.
For another perspective, principal component analysis (Figure 3) helps to identify the directions of the variation in the results and marks possible correlations between samples and the analyzed compounds.So, as far as melatonin is concerned, the influence of varietal variability was clear.Very high correlations (r > 0.9) between ferulic, p-coumaric and caffeic acid could be observed.High correlations (r > 7) were presented by gallic, caffeic, and ferulic acids, while a medium correlation of melatonin and protocatechuic acid was registered (r = 0.610).The effects of yeasts on the chemical composition of wines have been intensively studied; numerous studies followed the influence of similar oenological products [37,38], but few studies focused on the variation in melatonin content.In general, samples with a high content of melatonin also show higher antioxidant activity, which confirms the results obtained in other studies [18].
Table 6.The contribution of each bioactive compound on total antioxidant capacity of Sauvignon blanc wines.
In the case of Aligoté + Fetească albă samples, compounds such as trans-resveratrol, catechin, gallic acid, p-coumaric acid, and caftaric acid showed a negative influence on the TEAC value, suggesting a lower antioxidant activity.On the other hand, melatonin followed by cis-resveratrol, ferulic acid, protocatechuic acid, caffeic and epicatechin, presented a positive contribution to the increase in the TEAC value, indicating a greater antioxidant activity.For the second category, wines with bâtonnage, cis-resveratrol, protocatechuic acid, caffeic acid showed a negative impact on the TEAC value, while melatonin > catechin > p-coumaric > gallic acid > ferulic acid > trans-resveratrol > caftaric acid showed a positive contribution on the TEAC value.
In Sauvignon blanc wines, the TEAC value was negatively influenced by the concentration of compounds such as gallic, protocatechuic, caftaric, caffeic, p-coumaric, and ferulic acids.Also, the higher positive contribution was evident for melatonin, followed by cis-resveratrol, epicatechin, catechin, and trans-resveratrol.After the administration of bâtonnage products, melatonin exhibited the greatest influence, followed by p-coumaric, protocatechuic, cis-resveratrol and ferulic acids.
Therefore, the hypothesis is confirmed that although it is found in very small proportions in wines, melatonin may have made the largest contribution to antioxidant activity in the analyzed samples.Similar results have been reported previously.According to Sunyer-Figueres [35], melatonin exhibits direct antioxidant action (by eliminating reactive oxygen species) and indirect (by decreasing oxidized glutathione and activating genes involved in the response to oxidative stress such as catalase, glutathione, glutaredoxin and thioredoxin).Also, the authors postulated that melatonin confers protection against ethanol stress.Melatonin may act synergistically with other wine antioxidants, resulting in an increased cytoprotective impact against oxidative stress [8,9].The results are in accordance with Vasquez et al. [39], confirming that melatonin manifests important anti-scavenging action on Saccharomyces cerevisiae yeast but, in the present study, the results showed a better efficiency when Saccharomyces cerevisiae yeast was inoculated in combination with non-Saccharomyces species.
There are numerous chemical, environmental, and methodological elements that can interact with phenolic acids in wine and may influence their contribution to total antioxidant activity.These interactions can also have positive or negative oxidative effects.Therefore, phenolic acids can form complexes with other wine components, such as proteins, metals, or other phenolic compounds.Consequently, phenolic acids have the ability to form complexes with other elements found in wine, including proteins, metals, and other phenolic compounds.These complexes have the potential to modify the phenolic acids' availability or reactivity, which might impact their antioxidant efficacy.These complexes may occasionally promote oxidation processes as opposed to inhibiting them.Phenolic acids' antioxidant activity can change depending on the pH and external factors like temperature and oxygen exposure.For instance, phenolic acids' ionization state and reactivity can change in response to pH changes, which might impact their capacity to scavenge free radicals and contribute hydrogen atoms or electrons.The assessment of total antioxidant activity may also be impacted by the analysis technique adopted [40].This might be due to side reactions such as the formation of coupling adducts with ABTS + by different phenolic acids or a pro-oxidation reaction.Variations in the reported effects of phenolic acids may result from various assays that capture different features of antioxidant capacity or are more sensitive to particular types of antioxidants [40].
The presented results show that the antioxidant action of melatonin is dependent on various factors, such as the variability of the variety, the chemical composition and the applied technology.These variations may have an impact on the interactions of melatonin in each grape variety.It is important to explore more about the distinct qualities of each wine variety, their individual compositions, and the ways in which melatonin interacts with those components to explain the variations in its contribution to antioxidant activity that have been found.
* Values are presented according to the producers' technical sheets.

Table 4 .
Bioactive compounds in Sauvignon blanc wine samples.

Table 5 .
The contribution of each bioactive compound on total antioxidant capacity of Aligoté + Fetească albă wines.