Changes in anthocyanin pigments, trans-resveratrol, and colorimetric characteristics of Fondillón wine and other “Monastrell” wines during the aging period

The studied red wines, which are Fondillón, reserva, crianza, and young, are prepared from exclusive “Monastrell” grapes, and protected under the Alicante Denomination of Origin (Alicante, Spain). The objectives of the present study were (i) to show the effect of aging period on the amount of anthocyanin pigments, trans-resveratrol, and the colorimetric characteristics (color intensity, color density, and tonality) of Fondillón and other Monastrell wines (young, crianza, and reserva) which have different aging periods, and (ii) to propose a new traceability method for the determination of the Fondillón originality. Identification of anthocyanin compounds and determination of trans-resveratrol content were performed using LC–MS/MS. Correlation analysis was performed among anthocyanin compounds and color parameters. A sharp decrease in the anthocyanins content was observed in Fondillón wine. Besides, statistically significant changes were found in all colorimetric parameters between young and Fondillón samples. A significant change from red hue to brick red/yellow hue was observed between the colors of young and Fondillón wines, respectively. A significant decrease in the content of trans-resveratrol was also observed as a result of the aging period of these “Monastrell” wines. In this study, anthocyanin compounds of Fondillón wine were identified for the first time and their effects on the colorimetric parameters of the wine were revealed. Consequently, the authenticity of Fondillón wines can be determined by monitoring of anthocyanin compounds and trans-resveratrol.


Introduction
Wine contains a variety of phenolic compounds, most of which originate from grapes. These phenolic compounds are responsible for many essential functions in wine, and one of them is that they can affect both color and flavor, especially in red wine, and so the overall color of red wine is derived from phenolics [1,2]. Anthocyanins (flavonoid phenols), which are one of the crucial phenolic compounds for wine color, play a relevant role during wine aging and storage, mainly leading to wine browning [3]. They are mainly synthesized and accumulated into the grape skin through the binding of anthocyanidins and sugar moiety [4]. Secondly, a non-flavonoid phenol, a stilbene derivative, resveratrol is also another phenolic compound that can affect the color and functionality of red wine [1]. Resveratrol has two isomerization types in its free and glycoside forms that are -trans and -cis. While trans-resveratrol forms naturally in the grape skin as its glycoside, named as piceid [5], conversion from the trans-into the cis form can happen as a consequence of the isomerase activity during yeast fermentation [6].
Color is also one of the important factors affecting the organoleptic characteristics of wines and can also be affected by a number of parameters such as the grape type, grape composition, vinification techniques and various reactions that occur during wine storage [7]. Thus, several processing steps of red wine, particularly vinification and storage lead to changes continuously in color composition and sensory characteristics. These successive changes occurring in color composition are induced by reactivities of the phenolic substances such as anthocyanins and resveratrol extracted in fermentation [8]. Therefore, the effects of the vinification, storage, and aging processes of special wines on the organoleptic properties can be examined by analyzing the trans-resveratrol and anthocyanins composition of wine. In addition, conducting studies on the colorimetric parameters of wine and the compositions of anthocyanins and transresveratrol that cause the wine color will contribute to wine authentication and traceability.
In the literature, some studies have been reported on the determination of the anthocyanin and/or resveratrol compositions and on the evaluation of colorimetric parameters of different wines, which have different originality and are produced under various fermentation (pre-and post-) conditions [1,[7][8][9][10][11][12][13]. According to the abovementioned studies, application of different fermentation processes may cause significant effects and changes on the composition and amount of phenolics and colorimetric parameters of wine. However, the effect of changes in the composition of anthocyanins and the amount of trans-resveratrol due to aging period and process on the colorimetric character of Fondillón wine has not been studied in the literature. Especially for wines that have a unique aging process, studies on examining the effect of aging on both the chemical composition of the wine and its organoleptic properties are still insufficient.
Fondillón is a naturally sweet red wine and can only be produced under the Alicante Protected Designation of Origin, Alicante PDO [14]. It is produced from overripe "Monastrell" grapes and bottled after an aging process of at least 10 years in oak barrels resulting in an alcoholic content of over 16%. Fondillón can be considered as one of the late harvest wines due to dehydration processed on wine [15]. Overripe grapes include a rich composition of volatile compounds because of producing naturally concentrated must. In addition, it has been reported that Fondillón wines at different times of aging contain different amounts of phenolic compounds [16]. While these changes in some phenolic compounds have been studied in Fondillón samples from different soleras, the amount of anthocyanin compounds and trans-resveratrol and their effects on wine color have not been studied.
Consequently, the objectives of the present study were (i) to determine the content and composition of coloring substances, anthocyanins and trans-resveratrol, in Fondillón and other "Monastrell" wines (young, crianza, and reserva) aged for different years in oak barrels; (ii) to show the relationship among these compounds and the wine colorimetric characteristics (color intensity, tonality, and density); and, (iii) to propose a new approach to the authentication of Fondillón wines by monitoring new chemical identifiers of wine aging.

Wine samples
Four wine samples with a different seasoned time (young (2020), crianza (2018), reserva (2017), and Fondillón (1988)) were kindly provided by Bodegas BOCOPA (Petrer, Alicante, Spain). All the samples were provided in triplicate (from three batches of the same kind of wine) and were taken immediately to the facilities of the Miguel Hernández University of Elche (Orihuela, Alicante, Spain). Alcohol percentage, volatile acidity and total acidity analyses were analyzed according to the methods of the Office International de la Vigne et du Vin [17]. The pH measurements of the samples were taken by pH-meter (XS Instruments, 60 + DHS, Italy).

Chromatic characteristics of wine
The chromatic characteristics: (i) tonality, (ii) color intensity, and (iii) color density in the wine samples were analyzed according to the procedure described by [18]. Absorbance measurements were performed using a ThermoSpectronic Heios spectrophotometer (Cambridge, England). Color of wine samples was determined by measuring the absorbance at 420 nm (yellow components), 520 nm (red components), and 620 nm (blue components); color intensity (IC), tonality (T), color density (D), and brilliance of red color (dA) were calculated with the following formulas: The Glories color index percentages of yellow (Y %), red (R %), and blue (B %) of the wines was calculated IC = A420 + A520 + A620 dA (%) = (A520−(A420 − A620)∕2)∕A520 * 100 using the data obtained at 420, 520, and 620 nm [19]. All samples were filtered through a 0.45 µm pore size membrane filter before analysis and the wine color analyses were performed in triplicate.

Anthocyanins and polyphenol composition (LC-MS/ MS)
Wine samples were diluted (1:1, w/v) by addition of extractant consisting of methanol/water/formic acid (80:19.9:0.1, v/v). Then, diluted wine samples were filtered through a 0.45 µm pore size membrane filter before injection in the LC-MS/MS system. Anthocyanins and trans-resveratrol composition analyses were carried out on a Shimadzu LC-MS/MS 8050 triple quadrupole mass spectrometer equipped with an electrospray ionization (ESI) as source operating in negative and positive modes and a Shimadzu High Performance Liquid Chromatography (HPLC) system (Shimadzu, Kyoto, Japan). The HPLC equipment was used with a photodiode array detector (PDA) SPD-M40 (Shimadzu, Kyoto, Japan). The equipment system includes a binary pump, degasser and autosampler. The column used in this study was a Mediterranea SEA18 (100 mm × 2.1 mm, 2.2 µm) (Teknokroma, Barcelona, Spain) maintained at 50 ºC. The method of LC-MS/MS analysis was performed according to the procedure described by Wojdyło (2018) with a slight modification [20]. The mobile phase consisted of two solvents: (i) Solvent A, water/formic acid (99.9:0.1, v/v) and (ii) Solvent B, acetonitrile/formic acid (99.9:0.1, v/v). Anthocyanin compounds were eluted as following conditions: 0.4 mL min −1 flow rate and 30 ºC, isocratic conditions for 1 min with 99% A, from 1 to 15 min linear gradient of 1-40% acetonitrile with 0.1% formic acid (B), solvent B was increased to 100%, between 15 and 23 min, then returned to initial conditions of 99% A in 2 min, and isocratic conditions with 99% of 1% aqueous formic acid for 5 min followed by washing and reconditioning the column. The sample volume injected was 10 µL. The ultraviolet visible (UV-visible) spectra were scanned from 200 to 600 nm for all peaks. The analysis was performed in triplicate for each sample. The identification of anthocyanins and resveratrol was acquired using authentic standards and comparing the retention times and UV-visible spectra with those found in the literature. In addition, concentrations of the individual compounds were quantified using external calibration with the proper standard compounds. The stock solution of all individual standards was prepared with the proper extractant in the range of concentrations 0.1, 0.3, 0.5, 0.8, and 1.0 mg L −1 . Afterward, an aliquot of the prepared solution was injected (10 µL) into the HPLC apparatus.

Statistical analysis
All experiments were carried out in triplicate and results were expressed as mean ± standard deviation (SD). Statistical analysis was performed using XLSTAT Premium 2016 (Addinsoft, New York, NY). One-way analysis of variance (ANOVA) test was used to examine the significant differences in chromatic characteristics, anthocyanins, and polyphenol compounds of the wine samples. Then, Tukey's multiple range tests were applied for the post-hoc test, where the effect is commonly considered significant if the resulting p-value is below 0.05. A statistical analysis was also conducted to determine if there was a correlation among chromatic parameters of wines and the contents of anthocyanins and trans-resveratrol; correlation coefficients of the analysis were reported.

The physicochemical parameters of wines
The physicochemical quality parameters and season of wine samples are presented in Table 1. All these red wine samples were produced using the same type of Monastrell grapes with the application of different aging periods defined by the Alicante Protected Designation of Origin, PDO [21]. According to Alicante PDO, crianza wines must have a minimum aging period of 24 months, of which they must remain at least 6 months in oak barrels with a maximum capacity of 330 L. The other wine, reserva must have a minimum aging period of 36 months in total, including at least 12 months in oak barrels with a maximum capacity of 330 L and the rest of this period in the bottle. For the last wine, Fondillón must be produced from overripe Monastrell grapes, have at least ten years of aging time in oak barrels and acquire a minimum alcohol content 16% v/v.
As can be seen in Table 1, and depending on the aging periods, the main characteristics of these wine samples are: (i) young wine sample was prepared using exclusively Monastrell grapes from the 2020 season, with 15% alcohol, total acidity (5.2 g L −1 tartaric acid), volatile acidity (0.7 g L −1 acetic acid), and pH 4.03; (ii) aged/crianza wine was produced using exclusively Monastrell grapes from the 2018 season, with 15% alcohol, total acidity (5.3 g L −1 tartaric acid), volatile acidity (0.6 g L −1 acetic acid) and pH 3.79; (iii) reserva wine was prepared using exclusively Monastrell grapes from the 2017 season, with 15% alcohol, total acidity (6.1 g L −1 tartaric acid), volatile acidity (0.6 g L −1 acetic acid), and pH 3.77; and, (iv) Fondillón prepared exclusively from overripe Monastrell grapes from the 1988 season, with 16% alcohol, total acidity (5.9 g L −1 tartaric acid) and volatile acidity (0.8 g L −1 acetic acid).

Identification of anthocyanin and stilbene compounds of "Monastrell" wines
The anthocyanins and trans-resveratrol (trans-stilbene) were detected using positive mode, in which flavylium cations are produced under acidic conditions, whereas the negative mode is frequently selected for carboxylic acids and uncharged flavonoids that are found as deprotonated [M-H] − [20]. The anthocyanin compounds which were identified and quantified in the wine samples are presented in Table 2. Ten, eight, seven, and three anthocyanin compounds could be identified for young, crianza, reserva, and Fondillón wine samples, respectively. Anthocyanidin is the term for the simple flavonoid ring system and there are five basic anthocyanidins in wine: malvidin, delphinidin, cyanidin, peonidin, and petunidin; besides, "anthocyanins" refer to compounds including a glycoside [2]. In the wine samples, pelargonidin and five basic anthocyanidin structures which are cyanidin, peonidin, delphinidin, petunidin, and malvidin were identified with a molecular product ion at m/z values of 271, 287, 301, 303, 317, and 331, respectively, indicating the loss of sugar moiety (such as glucose). Six monoglucosides were found in the samples by presence of the pro-  [2,20,22]. The reason why this compound is found in trace amounts in one sample is because the "Monastrell" grape belongs to the Vitis vinifera species.
Apart from glucoside forms of the compounds, 3-O-rutinoside derivatives of cyanidin (compound 5, (M + H) + at m/z 595.10) and pelargonidin (compound 8, (M + H) + at m/z 579.20) compounds were also found in the samples by the presence of the specific protonated molecular ion and representing a loss of 308 Da fragment corresponding to the loss of rhamnosyl-glucosyl moiety [23]. Malvidin-3-O-galactoside (compound 9, (M + H) + at m/z 493.20, t R = 8.59 min) was also identified in the wines by comparison of standard molecular ion and the fragment (M + -162) related with the loss of galactose moiety [24]. Furthermore, t-resveratrol compound (compound 1, (M + H) + at m/z 229, t R = 13.07 min) was detected in the samples compared with standard and a protonated molecular ion fragment which corresponded to free resveratrol. As a conclusion, several anthocyanin and trans-resveratrol compounds of Fondillón and other young ''Monastrell'' wines have been identified quantitatively using LC-MS/MS.

Comparison of anthocyanin and stilbene compounds of wines
The profile and amount of anthocyanin compounds in wine samples largely depend on the aging process in both oak barrels and/or bottles. In young "Monastrell" wine, while the amount of seven compounds could be successfully determined, the rest of the anthocyanins (pelargonidin-3,5-di-O-glucoside, pelargonidin-3-O-glucoside, and cyanidin-3-Orutinoside) were found only in trace amounts. As can be seen from Table 2, for the crianza sample, the amount of seven same compounds as in the young wine could be quantified but other two compounds (pelargonidin-3,5-di-O-glucoside, and pelargonidin-3-O-glucoside) were not detected. It was seen that the anthocyanin compounds detected in almost all wine samples were the compounds (peonidin-3-Oglucoside, malvidin-3-O-glucoside, and malvidin-3-Ogalactoside) found with the highest amount (0.306, 0.667, and 0.661 mg L −1 , respectively) in the first wine sample. These compounds were found to be the most abundant anthocyanins in the "Monastrell'' wine samples under study. Malvidin derivatives compounds are also found as one of the most abundant compounds in several wine samples in the scientific literature [11,20]. On the other hand, apart from these components, delphinidin-3,5-di-O-glucoside, cyanidin-3,5-di-O-glucoside, and cyanidin-3-O-arabinoside compounds were analyzed but could not be detected. If the total amounts of anthocyanins (1.877, 1.214, 0.735, and 0.005 mg L −1 of young, crianza, reserva and Fondillón wines, respectively) are considered as affected by the aging period, a decreasing trend was observed as the wine aged. Considering all the above, these experimental results clearly showed that as the aging period gets longer, the anthocyanin profile gets less complex, and their contents significantly decrease. In another study, it has been also reported that young wine aged less than six months had more anthocyanin content than more aged wine (more than two years in oak barrels) [2]. Similar results have been noted in other studies by observing the losses of total pigments and free anthocyanins along the aging period of red wines [25,26]. During the red wine aging process, phenolics undergo several reactions, which are dependent on temperature, degree of oxidation, time, anthocyanate to tannin ratio, etc. One of these reactions, condensation can occur between anthocyanins and tannins forming tannin-anthocyanate complexes. These condensation products could lead to change in color towards tile red hues. In addition, anthocyanins can be degraded because of oxidation with formation of yellow or colorless degradation compounds [27,28]. In addition to these reactions, a series of mechanisms might be linked with such changes, such as anthocyanins precipitation with polysaccharides, proteins, or condensed tannins, adsorption by yeast, formation of more complex and further stable anthocyanin-derived pigments, named as several pyranoanthocyanins, polymeric anthocyanins formed by condensation between anthocyanin and/or flavan-3-ols, and their progressive derivatives [29][30][31][32].
Another compound, one of a major stilbene phytoalexin, trans-resveratrol was also determined for young, crianza, and reserva samples with a content of 0.146, 0.619, and 0.093 mg L −1 , respectively. However, this compound was not found in the Fondillón sample. Considering the changes from the young sample to that of Fondillón, trans-resveratrol content significantly decreased during the aging process. The findings observed in the loss of amount of trans-resveratrol leading to a full disappearance at the end of aging and storage agrees with previous research in the literature [33][34][35]. This is most likely due to isomerization of transresveratrol to cis-resveratrol by enzymatic transformation, one of the main processes during vinification [6,36] and in oak barrel aging [34] that can continue during the final bottle storage [35]. This disappearance of trans-resveratrol in Fondillón wine also agreed with previous studies conducted with long-aged (more than 50 years) Fondillón wine samples. Free trans-or cis-forms could not be detected, whereas only cis-piceid forms were found [16]. On the other hand, there is a significant increase in the amount of trans-resveratrol between young (0.146 mg L −1 ) and crianza (0.619 mg L −1 ) wine samples. Normally, trans-resveratrol occurs naturally in the grape skin as its glycoside form (piceid). However, during alcoholic fermentation, the β-glycosidase activity of yeast also releases trans-resveratrol [5,37]; this may be the reason for the increase in the content of this compound between two samples.

Chromatic characteristics of wines
Color intensity (CI), tonality (T), color density (D), yellow (%), red (%), blue (%) percentages of wine samples were determined using spectrophotometry. The values of all chromatic parameters of wine samples are given in Table 3. It can be seen that the pigments composition reflects absolutely the chromatic features of all wine samples. Compared to other wine samples (young, crianza, and reserva), Fondillón showed statistically a clear difference in yellow and red pigment category. It has the highest value for the percent of yellow shade (52.1%), while has the lowest percent for the red (35.3%) and blue (12.6%) pigments among the other wines. Besides, for all other wines (young, crianza, and reserva), the percentage of yellow pigment was below 37%, while the Table 3 Chromatic characteristics, determined by Glories method, of the "Monastrell" wine samples under study † , ***Significant at p < 0.001. ‡ Values (mean of three replications) followed by the same letter, within the same column, were not significantly different (p > 0.05), according to Tukey's least significant difference test. ¶ Y: yellow color, R: red color, and B: blue color. A420: absorbance at 420 nm, A520: absorbance at 520 nm, A620: absorbance at 620 nm, dA: brillance of red color Wine type 12.6 ± 0.03 d red pigment percent was above 45%, therefore the shade of these wines was red, while for Fondillón wine the shade was brownish. The reason for the changes in yellow and red pigments for aging of red wines could be due to the shift from monomeric (free anthocyanins) to polymeric anthocyanins leading to an increase in yellow percentage [38,39]. In this situation, some reactions among malvidin-3-glucoside and catechin and acetaldehyde might give an increase in the content of polymeric contents [8]. The formation of anthocyanin-derived pigments during aging period seems to lead to change in the color of wine from the purple-red pigment of young red wine to a brick-red pigment with a characteristic color of aged wines [40] Furthermore, in addition to the phenolics originating from the grape, the components from the oak barrel also cause significant changes in wine. In this way, condensation reactions take place between phenolic substances such as anthocyanidins, tannins sourced from the wine and other compounds extracted from the stave of cask, represented by hydrolysable tannins [12]. Considering the anthocyanins analysis, change in the percent of red and yellow pigments was also found consistent with a decline in the amount of total monomeric (free) anthocyanins and free malvidin-3-glucoside of Fondillón wine. In another study, similar observations in the change in dispersion of yellow and red pigments were also reported for the wine aged for a long period in oak barrels [12].
On the other hand, a slight change was observed in the blue pigment of Fondillón (12.6%) compared to other less aged wines (young, 17.3%; crianza, 17.1%; and reserva, 18%). A decrease in the blue hue might have resulted from the formation of new compounds during aging leading to change in the shift of red pigment to yellow pigment of the samples. Anthocyanins are also responsible for the blue hue of red wines. The reaction among free anthocyanins and some of yeast by-products, such as acetaldehyde [41], pyruvic acid [42], vinilcatechol [43], and vinylguaiacol [44], may lead to the formation of different group of stabilized pigments named as pyranoanthocyanins [40]. While most pyranoanthocyanins have yellow-orange color contributing to the tawny color (also decreasing blue color) shift related with wine aging, another resulting pyranoanthocyanin, named portisins provide blue color to the red wine [32,45,46]. Thus, the reason for the slight decrease observed in the percent of blue pigment compared to the change in other colors for Fondillón might be explained by the formation of these components (portisins). Considering all the abovedescribed findings, it can be stated that the percentage of the pigments is decisive in the overall color of the wine. Since red pigments were dominant (̴ 46%) in the three less aged wine samples, their color was perceived as intense red; however, the total color of Fondillón wine was perceived as browny, not red, because the contribution of the yellow pigment (52.1%) was more dominant than red pigment (35.3%).
For the values of color intensity, young (7.95), crianza (7.87), and reserva (7.99) wines were found statistically different, while the results were observed very close to each other. On the other hand, Fondillón wine aged 34 years has the lowest value (5.21) among the wine samples, with a remarkable decrease in the color intensity. This is a result of having the lowest value of all the pigments percentage for Fondillón. Another characteristic, color density as defined by Sudraud (1958) [47] as the estimate of the contributions of yellow (420 nm) and red (520 nm) colors, exhibits a similar trend as that described for the color intensity values. The values of young, crianza, reserva, and Fondillón were calculated as 6.57, 6.52, 6.55, and 4.56, respectively. As reported above for CI, Fondillón was the wine sample with the lowest density value. The results showed that the color density and intensity values of wines decreased as affected by the aging period.
While the tonality values of the young samples (young, 0.80; crianza, 0.79; and reserva, 0.79) were found very close to each other, the tonality value of Fondillón (1.48) was statistically higher than the others. As wine tonality (yellow pigment (%)/red pigment (%)) values range between 0.6 and 1.0 the wines are characterized by red shade, while for values above 1.0 they have a brownish shade, which is a sign of age and/or level of oxidation [12]. For Fondillón wine, the aging process in the barrel can be visible by observing its tonality values (> 1.0) and changing in chromatic structure (yellow-orange pigments participate more than red pigments at the total wine color). Another characteristic in the colorimetric, the brilliance of red color (dA) of Fondillón wine (43.9%) was also obtained statistically different and as having the lowest value among the young (78.9%), crianza (79%), and reserva (80.2%) wine samples. The brilliance of red wines is related to the shape of the spectrum. When the wine has a bright red, the maximum spectrum at 520 nm shows narrow and well defined. Once the maximum spectrum is a broad and flattened shape, wine has a deep red or brick red [48].

Correlation between colorimetric characteristics and phenolic compounds of wines
Pearson's correlation analysis was performed among the colorimetric characteristics (color intensity, tonality, density, yellow (%), red (%), and blue (%)), anthocyanin, and trans-resveratrol compounds of all wine samples. Correlation coefficient matrix of the analysis between the chromatic parameters and phenolic compounds (anthocyanins and trans-resveratrol) is given in Table 4. It was found a statistically significant (p < 0.05) positive relation observing high correlation coefficients among the anthocyanin compounds (malvidin-3-O-glucoside, malvidin-3-O-galactoside, petunidin-3-O-glucoside, and peonidin-3-O-glucoside). The Table 4 Pearson correlation coefficients matrix among the chromatic parameters and phenolic compounds found in the "Monastrell" wine samples under study † Values in bold font are statistically significant *, **, and *** at p relationship of these anthocyanin components with each other is because of the decrease in their amounts in the wine samples depending on the aging period. Another positive correlation was also obtained between delphinidin-3-O-glucoside and cyanidin-3-O-glucoside with a high correlation coefficient (r = 0.990). The difference in the analysis of these two anthocyanins is that the amount of these compounds was less than the other anthocyanin compounds in the first two wines (young and crianza) and then, they disappeared in the other aged (reserva and Fondillón) wines. Considering all above results, it can be deduced that all free monoanthocyanin compounds underwent a series of reactions, and they disappeared or present with a very low amount in the Fondillón wine sample. A significant positive or negative correlation was found among all colorimetric characteristics of wine samples. As can be seen from the table, a high negative correlation between yellow-red pigment (r = − 0.998) and yellow-blue pigment (r = − 0.991). On the other part, red and blue pigments of the samples showed a highly significant and positive correlation (r = 0.980). These findings in correlation are compatible with the above results changing in yellow (%) and red (%) colors of the wine samples as aging period increases. Red and blue colors contribute to the color intensity with a positive correlation, whereas yellow color shows a high negative correlation between the total color intensity of the samples. The reason is that the decrease in the absorbance wavelength of yellow color (A 420 , 2.72) in Fondillón wine was not as sharp as observed with the red (A 520 , 1.84) and blue (A 620, 0.66) pigments. The results could be explained by comparing with previous findings (in 3.2 and 3.3 chapters) that occurrence of some reactions in the anthocyanins compounds led to change in the color from red hue to brick red-yellow hue. While a positive correlation (r = 1.00) was also obtained between yellow (%) pigment and tonality value, a high negative relation (r = − 0.99) for density value was found between these variables. A statistically significant relationship was not found among (i) colorimetric characteristics and trans-resveratrol or (ii) anthocyanin compounds and trans-resveratrol. This may be due to the increase in the content of trans-resveratrol in crianza wine by yeast fermentation. However, it has been shown that there is a negative relationship between the aging period and trans-resveratrol, although it cannot be found statistically. Though small amounts of resveratrol were also found in the young sample, it might influence the color intensity, red color (%) and pigmentation reactions in the wine. Ulrih et al. [49] reported that addition of trans-resveratrol to the red wine samples increased the intensity of color and portions of red color and then initiated the formation of new copigments causing an increase in color intensity. In another study, similar results were also reported that color density of the wine samples were obtained with a high value by increasing the amount of trans-resveratrol fortification. Thus, t-resveratrol was associated with a high ratio of red to yellow-brown colored pigments [50].
As a conclusion, when the content of anthocyanin compounds and colorimetric characteristics of the samples were evaluated together, it was revealed that there was a relationship between these two variables of wine samples. It can be noted that aging period and conditions have an effect on the phenolic compounds of wine samples and by the effect of the changes in these phenolic components, lead to some transformations in organoleptic properties of wine samples. In the literature, there is no study on the identification and determination of anthocyanin compounds of Fondillón wine. Furthermore, there is no comprehensive "Monastrell" wine study on evaluating aging time, anthocyanins profile, transresveratrol, and colorimetric properties together.

Conclusion
In the present study, the profile and content of anthocyanin compounds were determined in Fondillón and other wines (young, crianza, and reserva) produced from "Monastrell" grapes. Malvidin-3-O-glucoside, malvidin-3-O-galactoside, and peonidin-3-O-glucoside were found to be the most abundant anthocyanins. A high negative correlation was obtained between the content of anthocyanin compounds and the aging period of wine samples. As the aging time of wine samples increased, a decrease in the content of all anthocyanin compounds was observed. Furthermore, colorimetric parameters of the wine samples have been affected by the aging period of wines. Fondillón has the lowest red color (%) content, whereas it has the highest level of yellow pigment (%) of the studied wine samples. It was revealed that a long period of fermentation in oak barrels facilitated a series of reactions among anthocyanin and other compounds resulting in a new co-pigmentation. trans-Resveratrol, one of major stilbene compounds in red wine, was not detected in Fondillón wine due to the isomerization reaction transforming cis form by yeast fermentation into the trans isomer. As a result, the aging experienced by Fondillón caused a significant change from the initial red/purple color to the final yellow/brick-red color, and to a decrease in the contents of free mono-and di-anthocyanins and trans-resveratrol compounds. Besides, this study revealed that anthocyanin and trans-resveratrol compounds can be used as a quality control marker of Fondillón authentication.