Violeta+Nasto

**Violeta Nasto Log**

 * An Overview of the Various Compounds Contributing to Aroma, Flavor, and Color in Red Wine**


 * Introduction**

Whenever one is about to enjoy drinking a glass of red wine, they should try to consider thinking where the aroma and flavor of that glass of red wine is derived from or even how the rich red color has gotten its intensity shown in that glass of wine one is experiencing. The compounds involved in these three properties of wine and the different factors that can have an effect has been shown to be numerous and complex. Significant information has been learned, even though further studies are necessary to conclude the many compounds found in red wine, ways to analyze those compounds, and the factors that can cause changes to the properties of wine such as the aroma, flavor, and its red color.


 * Analytical Methods**

Much information, especially of the compounds present in wines, has been found with use of analytical tools. Even the use of analytical tools has been found to be challenges since there over a thousand volatile compounds under many chemical classes that are present in wines (14). Some of these compounds include esters, alcohols, terpenes, and even sulfur compounds with different concentrations in each component, making it harder and presenting challenges in analyzing each component or the number of components present (14). Since these components are volatile; however, gas chromatography has been used for analysis. Even in this kind of analysis though, it is very important to have proper sample preparation. Techniques in isolation these volatiles has been to use distillation or solvent extraction, but again, these can show to be methods that involve multi-step procedures and lead to analyte losses and other complications (14).

An approach that has been reviewed for this kind of analysis is the headspace technique. In this kind of technique, a gas-tight syringe is used to take an aliquot of the gas in equilibrium above the solution, or static headspace or the headspace is swept towards a sorbent that traps and preconcentrates the dynamic headspace (14). From there, the headspace aliquot is directly injected into the gas chromatographer for the headspace analysis (14).

Solid-phase microextraction or SPME is an alternative technique that is faster and involves no solvents for the isolation of analytics in a sample matrix. SPME extractions can be completed in about thirty minutes or even less when compared to the other solvent extraction procedures which would take several hours to complete (14). This technique has been widely used for the analysis of aromas in wine because it can be used with liquid, gaseous or solid samples and therefore excluding the need for solvent extraction (14). The SPME assembly contains a needle that has a retractable fiber coated with a polymeric sorbent material. This fiber is pierced through the septum of the vial that contains the sample, so this exposure to the sample headspace, concentrates the volatiles on the polymer. Then the SPME assembly is transferred to the heated GC injector, where furthermore, the carrier gas exposes the fiber and the volatiles are desorbed (14). Furthermore, there are a variety of SPME fibers which are coated with different polymeric materials, which lets it allow sorbing of different classes of volatiles and influences the selectivity of the extraction (14). As to any kind of technique though, there are numerous variables that would influence the effectiveness and accuracy of the technique. In SPME, the factors are the amount of time that the fiber is exposed, the temperature of the sample for analysis, and furthermore, in case of liquid samples, the pH, ionic strength, and the type of matrix composition that may be present are variables involved (14). Overall, it is necessary that the variables are optimized in order to enhance the sensitivity, accuracy, and reproducibility of the method for each of the analyte and matrix type (14). For example, in one study where the SPME method was used, matrix effects were noticed when the study was analyzing soaks that were obtained by the extraction of a high number of cork stoppers (20). In order for optimization of obtaining reliable and repeatable quantitative data, the standard addition calibration technique was applied and evaluated in real samples of wines (20).

In grapes and wines, matrix effects can occur during HS-SPME analysis, so there has been ways found to avoid these matrix effects. SIDA, stable isotope dilution analysis, has been a technique used to avoid the effects. In this technique, the wine samples are spiked with stable, isotopically labeled IS, internal standards, which are matched to the analyte of interest and therefore any interactions with the matrix would be compared with both analyte and the IS because of the similarity in the chemical structure (14). Moreover, the spiked samples can be analyzed by SPME-GC coupled to a mass spectrometer detector. The response ratio of analyte to internal standard is used to quantify the analyte concentrations in the sample by using calibration curves for each compound of interest that have been previously determined (14).

The SPME technique has been shown to be successful and have many applications with or without SIDA for analysis in grapes and wines. For example, in a study that used the SPME-GC-MS method combined with SIDA, it successfully quantified thirty-one different fermentation derived compounds found in wine such as fatty acids, alcohols, acetates, etc. by using twenty-nine different deuterated compounds as internal standards. The precision (<5%) was found to be very good for all compounds by using the SIDA technique (16, 18). Furthermore, another study was conducted using this same method, HS-SPME SIDA to get information about levels of aroma compounds which are newly identified in wines such as 13-norisoprenoid aroma compound and (E)-1-(2,3,6-trimethylphenyl)buta-1,3-diene (TPB) (16, 5). This method allowed for fast quantization at low levels of concentrations and be able to compare levels in different varieties. It also allowed monitoring the chemical changes that would occur during aging or processing of the wines (16, 5).


 * Aroma and Flavor**

There are many different components that play a role in the quality and more specifically on the aroma and flavor of wine. These components include not only the different compounds present in wine, but also factors that can affect the compositions of these compounds, as well as the overall amount of compounds found in wine. The many different compounds can come from grapes, Saccharomyces fermentation, microbes, and other processes such as oak barrels and bottle aging. Red wines are fermented with grape skins present, so many of the compounds can therefore come from the skins and in wine during fermentation and give complex aromas and flavor to wine (14).

Fermentation of red wine occurs in the following way. Red grapes are obtained and can be crushed. Saccharomyces is added to the whole red must acquired and it is fermented to get the red wine. From there, it can be pressed and let to settle or it can furthermore undergo secondary microbial fermentation, filtered, stored/aged in oak, or transferred to bottle (14).

Though sensory differences can vary in the aromas of different grapes, the overall compositions of the volatile components is similar. The largest percentage of the aroma composition in wine comes from the volatiles that are derived from fermentation (14). For example, there are esters formed during fermentation such as fatty acid ethyl esters and acetate esters (6). Ethyl acetate and isoamyl acetate are found to predominate, but actually many of the esters formed during fermentation are found not to contribute to overall flavor and aroma perception (6). Certain trace amounts such as terpenes and methoxypyrazines are found to have a larger contribution to the flavor of wine (6).

Terpenes are present in the skins of grapes and overall they give wines important floral and citrus characters. Levels of terpenes can be different depending on the maturation of grapes. Typically the levels of terpenes can increase as the grape matures (6). Also, other factors such as the fermentation process can change their levels because of the different acidic conditions. Storage can also have an affect on the terpenes as it can lead to the formation of new compounds. These compounds can furthermore contribute to different aroma qualities and intensities. Moreover, even climate and viticultural conditions can also have an effect on the terpene compositions (6).

Different types of methoxypyrazines can also contribute to the flavor and aroma of wine. Methoxypyrazines such as 3-isobutyl-2-methoxypyrazine (IBMP), 2-sec-butyl-3-methoxypyrazine (SBMP), and 3-isopropyl-2-methoxypyrazine (IPMP) have been identified (6). IBMP found in grapes obtains the aroma of bell peppers and leads to the sensory perception of the wine. SBMP and IPMP have levels that range depending on the variety, maturity, and growing conditions. For example, light exposure degrades IBMP and its levels decrease with grape maturation (6).

C13-Norisoprenoids, which are present in only trace levels, can also have an impact in the aroma properties of the wine. They are a diverse group of aroma compounds with very low sensory thresholds (6). Norisoprenoids are found to be produced from photochemical and thermally induced degradation of carotenoids found in the grapes (6). Their precursors are nonvolatile glycoside. It is hydrolysis occurring during fermentation and storage that releases these compounds and the contribution of their aroma in wine. As other compounds, climate, sunlight exposure, and maturity of grapes play a role on the levels of norisoprenoids found in red wine.

Other compounds obtained from the way wines are stored or processed can be found to contribute to aroma and flavor. Usually compounds from aging oak barrels are lactones and phenolic compounds. Wines that are stored in oak barrels for example contain the β-methyl-γ-Octalactone which gives the wine a “woody, oaky, coconut-like aroma” (14). Moreover, some examples of compounds that give different flavors and aroma to wine are as follows. Glucose, fructose, glycerol, and ethanol can give wine a sweet taste. Tartaric acid gives wine a sour taste. Sodium chloride and potassium chloride contribute to the salty taste and catechin gives wine a bitter taste (14). The compounds that give aroma to wine include linalool and isoamyl acetate. Linalool gives a floral aroma, whereas isoamyl acetate gives a “banana-like” aroma (14).

Even enzymes can be factors in wines. When a study was conducted on addition of enzymes to wine; it was found that there was an overall increase in the extraction of phenolic compounds. More specifically, addition of pectoltic enzymes showed enrichment for red wine and maintained higher quantities during the time that wines were stored (15). Linking enzymes to polyphenols could even contribute to studies such as aiming to determine effects of polyphenolic extracts that may be beneficial in prevention of cardiovascular diseases (10).

Chemical and microbial oxidative reactions moreover can have an effect. They contribute significantly to the flavor of aged wines because compounds like acetaldehyde and acetic acid are formed, giving wines a nutty and vinegar aroma, respectively. Oxidative reactions need to be controlled though, otherwise, the concentrations of these compounds can increase, which would lead to an undesirable sensory impact (14).

It is hard to connect the taste to a particular molecule because many compounds have shown low detection thresholds. The nose can only detect very small amounts that are hard to be identified as compounds by chemistry equipments (4). It can be said that wine aroma and flavor can’t be fully understood or predicted because even though many compounds have been found to contribute, there can be interactions that are formed between odorants and other nonvolatile components such as proteins, polysaccharides, lipids, and polyphenols (6). These interactions can furthermore change the concentration of the aroma compound during fermentation and processing. Polyphenols and tannins, which give a bitter taste and astringency, are also part of the nonvolatile matrix components of grapes and wines that can form these interactions. They can interact with the odorant compounds and alter the odorant volatility and aroma perception (6). Overall, the contents of phenolics in red wine again depend on the varieties of grape, on the maturations of the grapes, the producing areas, and even the different processes and wine ageing (7).


 * Color**

Controlling or predicting the final color in red wine also has not fully been understood. Red wines have very complex phenolic compositions which contain tannins, anthocyanins, and polymer pigments. These compositions can be affected by the winemaking techniques and changes of these phenolic compositions can even occur over the years (2). Pigmented molecules continually change from the grape, during crush and fermentation, the pressing, and even afterwards in the barrel during aging. For example, color can be obtained from tannins. The color extracted from tannins; however, is not always the same. The tannins from grape skins are usually extracted early in fermentation and the extraction rate increases as the maceration time increases (9). The maximum extraction of color occurs right before the end of fermentation (1). This also occurs at high temperatures, but it does not necessarily mean that if fermented at higher temperatures, it would result with more color (1). The tannins tend to usually increase throughout.

In different types of processing as well, the levels can differ. A study on red Merlot wine, showed that levels of polyphenols, tannins, and polymer pigments can change when exposed to heat-treated and non-treated oak wood chips. Exposing the wine to the mixed wood chips, allowed for the extension of phenolic compounds from wood to be faster and there were more phenolic compounds passed into wine when compared to the heated wood. In another study, a different process to stabilize the color proved to be an alternative to aging in barrels. The use of micro-oxygenation, enological tannins, and roasted oak woods showed to be “feasible and low-cost”. Studies done on micro-oxidation of wines do not indicate any changes or modify any enological parameters. Their results showed that the overall phenolic compositions were similar in all wines (16).

Polyphenols such as anthocyanins are also significant contributors to color. Malvidin-3-glucoside is a specific anthocyanin found in red wine (12). Anthocyanins; however, tend not to stay stable for a long time, so usually the anthocyanins “must evolve towards, new visible light absorbing, chemical chromophores” (3). Anthocyanins were found to be linked to tannins. Anthocyanins and condensed tannins are found to be the two most abundant classes of polyphenols found in grapes (8). Anthocyanins and tannins tend to condense in wine to form secondary wine pigments (1). The actual wine pigments are generated when anthocyanins combine with tannins, not the grape pigments that are usually found in the grape skins or seeds or in the yeasts. Specifically, it was found in the pigments of red wine, that usually the anthocyanin monomer is bound to the tannin monomer. SO2, however, was found by a study to have an effect on the level of anthocyanins. It was found that wines with lower concentration of SO2 had a large decrease in the monomeric anthocyanins. In one study, the extraction and ionization of total and individual anthocyanins were always higher when only SO2 was used (17). Depending on the SO2 concentration, polypehenol chemistry can be regulated by the formation of polymeric pigments and the change in tannin structure (19). Reactions formed are influenced by the relative amounts of anthocyanins and tannins, dissolved oxygen, pH, which controls reactivity, and even in presence of yeast metabolites. Rapid polymerization between anthocyanins and tannins, mediated by acetaldehyde, increases color intensity and stability (11). Moreover, anthocyanin-derived pigments were found by a study to be depended on the yeast strain during fermentation and that dependence can affect the preservation of wine color (13). These pigments allow the red wine to stay red over the years. This shows that there are even pigmented polymers that contribute to the color in red wine. And furthermore, the color tends to grow as wine ages (1).

Experiments have been conducted of nine winemaking technologies with addition of tannins in red wine. The experiments showed that the adding tannins obtained from the grape seeds positively affect the phenolic content and after pressing, the addition of tannins showed the higher total phenolic concentration in wines (2). The total phenolic content of one year old aged wines was found to be significantly higher than those young wines for all winemaking procedures (2). Also, when looking at aged wines, factors that play role in its composition and properties are the procedures the wine was made and the reactions that took place during the aging time. Wines that have the richest total anthocyanins are the ones produced traditionally. The addition of tannins and the heating of must-wine resulted in the highest content in the monomeric anthocyanins (2).

A study that worked on prediction of wine color from the phenolic profiles of fifty-five different red grape samples, showed a good relation between the anthocyanins from wine and the anthocyanins from the grapes. It was found that the prediction lead to the same precision as from the direct relation with the grape anthocyanins. The same relation was found between grape anthocyanins and total wine color (8).


 * Conclusion**

There are a high number of compounds found in red wine. Many have been identified and analyzed with various analytical methods. Such techniques include distillation, solvent extraction, and more efficiently, solid-phase microextraction (SPME). Even though various methods have been found to identify, quantify, and analyze these compounds, there are always variables that present challenges in the use of these methods and overall do not allow for every compound, its concentrations, and composition to be identified in wines.

Several compounds that are involved in aroma and flavor of red wine are volatiles derived from fermentation. Compounds can also come from the skins of the grapes. Such compounds include terpenes and methoxypyrazines. Other compounds are derived from the process of wine-making, such as lactones and phenolic compounds. Compounds that contribute to the color of red wines include tannins, anthocyanins, and polymer pigments. It is important to note that even though compounds have been identified to give aroma, flavor, and color to red wines, many of these compounds can be in different levels due to various factors such as enzymes, chemical and microbial oxidative reactions, maturation of grapes, processing, and furthermore, aging. Due to these factors and more, it has been found challenging to identify all of the compounds found in wine or control and predict the final aroma, flavor, and color of a desired red wine.


 * References:**

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