Formation of Anthocyanins

Schwarz, M., Wabnitz, T.C., and Winterhalter, R, Pathway leading to the formation of anthocyanin-vinylphenol adducts and related pigments in red wines, J. Agric. Food Chem., 51, 3682, 2003. 

Asen, S., Eactors affecting formation of anthocyanin-flavonol co-pigment complexes and their importance on flower color. Plant Physiol, 47, 20, 1971. 

Duenas, M., Fulcrand, H., and Cheynier, V., Formation of anthocyanin-flavanol adducts in model solutions. Anal. Chem. Acta, 563, 15, 2006. 

The spectra were recorded in the positive-ion mode in the range of m/z 120-1 500. Some chromatograms illustrating the effect of aldehydes on the interaction of mv3gl and B2-3 -gallate are shown in Fig. 2.120. The chromatograms demonstrate that different aldehydes influence differently the formation of anthocyanin-flavanol pigments. The results of HPLC-MS measurements are compiled in Table 2.94. Because free aldehydes display an unpleasant aroma in Port wine these reactions may improve the quality of wines and contribute to the colour formation [266],... 

The R locus determines the presence (R) or absence (r) of anthocyanins in the seed coat. R is required (with i and T) to produce black seed [10]. However the identity of the gene product encoded by this locus has not been reported. Todd and Vodkin [25] have demonstrated that brown seed coats (r) contain proanthocyanidin (PAs) and black seed coats (R) contain anthocyanins in addition to PAs and suggested that R acts subsequent to the formation of leucoanthocyanidin but previous to the formation of anthocyanins. UDP-glucose flavonoid 3-0-glucosyltransferase (UF3GT) should be considered a candidate gene of the R locus but its identiflcation has not yet been reported. 

FIGURE 3.2 General phenylpropanoid and flavonoid bios5mthetic pathways. The B-ring hydroxylation steps are not shown. For formation of anthocyanins from leucoanthocyanidins two routes are represented a simplified scheme via the anthocyanidin (pelargonidin) and the likely in vivo route via the pseudobase. Enzyme abbreviations are defined in the text and in Table 3.1. 

Finally, reactions of flavonoid and nonflavonoid precursors are affected by other parameters like pH, temperature, presence of metal catalysts, etc. In particular, pH values determine the relative nucleophilic and electrophilic characters of both anthocyanins and flavanols. Studies performed in model solutions showed that acetaldehyde-mediated condensation is faster at pH 2.2 than at pH 4 and limited by the rate of aldehyde protonation. The formation of flavanol-anthocyanin adducts was also limited by the rate of proanthocyanidin cleavage, which was shown to take place at pH 3.2, but not at pH 3.8. Nucleophilic addition of anthocyanins was faster at pH 3.4 than at pH 1.7, but still took place at pH values much lower than those encountered in wine, as evidenced by the formation of anthocyanin-caffeoyltartaric acid adducts, methylmethine anthocyanin-flavanol adducts,and flavanol-anthocyanin adducts. The formation of pyranoanthocyanins requiring the flavylium cation was faster under more acidic conditions, as expected, but took place in the whole wine pH range. Thus, the availability of either the flavylium or the hemiketal form does not seem to limit any of the anthocyanin reactions. 

Vivar-Quintana, A.M. et al.. Formation of anthocyanin-derived pigments in experimental red wines. Food Sci. Technol. Int. 5, 347, 1999. 

Halbwirth H, Martens S, Wienand U, Forkmann G, Stich K. 2003. Biochemical formation of anthocyanins in silk tissue of Zea mays. Plant Sci 164 489-495. 

Aldehyde-tannin and aldehyde-anthocyanin condensation reactions result in polymer formation (Figure 1). These polymers may be responsible for haze formation in wine and the polymers may eventually precipitate out of solution (26). The polymerized tannins have different flavor properties than the monomeric starting units (21-29) and formation of anthocyanin polymers affects wine color. In addition, these reactions may result in a reduction of aldehyde flavors in the wine. These condensation reactions are discussed more fully in other chapters of this volume. The formation of strong covalent bonds between the aldehyde and the tannin or anthocyanin makes recovery of the bound aldehydes difficult. 

Adducts of malvidin-3-glucoside with vinylcatechin, vinylepicatechin or vinyidiepicatechin (procyanidin B2), reported in model solutions containing malvidin-3-glucoside, acetaldehyde and the respective flavan-3-ol (Francia-Aricha et al., 1997) (Fig. 3, (b)), have also been detected in wine fractions (Mateus et al., 2002a, b Atanasova et al., 2002). In the latter work, the authors showed, via thiolysis, diat both proanthocyanidins and monomeric flavanols are involved in the formation of anthocyanin-vinylflavanol and anthocyanin-ediyl-flavanol pigments induced by acetaldehyde. 

Anthocyanin-pyruvic acid adducts are known to be more abundant in Port wines than in red table wines, as seen from previous analysis in our laboratories (data not shown) and as referred by other authors (77). This feature may be related to the higher levels of pyruvic acid expected in fortified wines as a result of a shortened fermentation. In fact, when wine spirit is added in order to stop fermentation, the pyruvic acid concentration is expected to be higher than when the fermentation is allowed to go to dryness. Effectively, the pyruvic acid excreted by the yeast at the beginning of the fermentation is further used in the yeast metabolism (35). Therefore, could favor the formation of anthocyanin-pyruvic acid adducts. 

The direct reaction between anthocyanins and hydroxycinnamic acids readily explains the formation of anthocyanin-vinylphenol-type adducts in red wines. At the time of writing, this is the only experimentally verified mechanism leading to the development of 4-vinylcatechol and 4-vinylsyringol pigments, as the free vinylphenols have neither been detected in wines nor was it possible to generate these compounds via enzymatic decarboxylation using yeasts commonly applied to red wine fermentation... 

In the formation of anthocyanin-proanthocyanidin polymers, it is hypothesized diat the anthocyanin position of the polymer depends upon whether an electrophilic position (C>4) or nucleophilic position (C-8 or C-6) of the anthocyanin becomes involved in the interflavonoid linkage. The former and latter reactions would involve a nucleophilic position (C-8 or C-6) and electrophilic position (C4 a carbonium ion derived by acidic cleavage) of proanthocyanidins leading to the formation of the A-T 14,26-27) and T-A type 12,14) polymers, respectively. 

In red wines, flavanols may also react directly with anthocyanins, leading to the formation of anthocyanin-flavanol and flavanol-anthocyanin adducts [283-285], The condensation may also occur mediated by aldehydes, such as acetaldehyde, furfural, HMF, benzaldehyde, and others, leading to the formation of alkyl-methine bridged adducts. These latter display the same /Lav in the visible region at around 540 nm, which is bathochromically shifted when compared with the original anthocyanin (/Lax 525 nm) [279, 280, 286-289], thereby contributing to the color change in red wine. 

A solution of 2- 3C-acetate (99% Sigma-Aldrich, Milwaukee, WI) was prepared as a 1 1 or 1 3 ratio of i C 12c isotopes in 1 mM stock solution. This labeled precursor was added to the Petri dishes on the fourth day. The Petri dishes were opened and the upper filters discarded. Excess water in the dishes was removed and replaced with 2 mL of 1 mM 2-i3C-acetate. The same procedure was repeated on the fifth and sixth day. At the end of the labeling period, the roots were excised, and 1 was extracted as described above. Labeling procedures were done under low-intensity green light to prevent the formation of anthocyanins by the sorghum roots. 

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