450 M. Monagas and B. Bartolom ́e
hydroxycinnamic acid and the anthocyanin without enzymatic support (Fig. 9A.3f).
The C-2 (nucleophilic) position of caffeic acid is initially linked to the C-4 (elec-
trophilic) position of malvidin-3-glucoside giving rise to an electron deficient inter-
mediate (carbenium ion) which is stabilized with the aromatic ring substituents
(electron donors) of the cinnamic acid moiety. The intermediate carbenium ion can
be intramolecularly trapped by the anthocyanin-OH group at C-5 forming a pyrane
ring. The final product is then formed byoxidation and decarboxylation of the inter-
mediate. Consequently, only cinnamic acids with electron-donor substituents, like
p-coumaric, ferulic, caffeicand sinapic acids, could be involved in the reaction in
order to stabilize the carbenium ion. Through this new mechanism it was possible
to explain why caffeic and sinapic acids, compounds for which there is no evi-
dence for an enzymatic decarboxylation bySaccharomyces cerevisiaeCD into their
respective 4-vinylphenols, can also give rise to hydroxyphenyl-pyranoanthocyanins
(Chatonnet et al. 1993).
Evidence in wine.Hydroxyphenyl-pyranoanthocyanins, firstly detected and iso-
lated from polymeric membranes employed for red wine microfiltration (Cameira
dos Santos et al. 1996), have been studied by UV-visible, mass and NMR spec-
trometry (Fulcrand et al. 1996). This study confirmed the presence of phenyl-pyrano
derivatives of malvidin-3-glucoside and malvidin-3-(6-p-coumaroyl)-glucoside
(Fulcrand et al. 1996). The corresponding phenyl-pyrano of the entire series of
anthocyanidin-3-glucosides and theirp-coumaroyl derivatives(with the exception of
cyanidin), malvidin-3-(6-acetyl)-glucoside and malvidin-3-(6-caffeoyl)-glucoside,
have been also reported in wines (Asenstorfer et al. 2001; Hayasaka and Asenstor-
fer 2002; Mateus et al. 2003a; Alcalde-Eon et al. 2004, 2006; Monagas et al. 2003;
Wang et al. 2003a; Pozo-Bay ́on et al. 2004; Boido et al. 2006).
Using a combination of mass spectrometry techniques (nano-ESI-MS/MS),
Hayasaka and Asenstorfer (2002) later identified a new series of hydroxyphenyl-
pyranoanthocyanins in red wine fractions, including the catechyl-, guaiacyl- and
siringyl-pyrano derivatives of malvidin-3-glucoside. Catechyl-pyranomalvidin-3-
glucoside (also called pinotin A) has been isolated fromVitis viniferacv Pinotage
red wine and characterized it by HPLC-ESI/MS and NMR (Schwarz et al. 2003a).
The presence of catechyl-pyrano derivatives of the entire series of anthocyanidin-
3-glucoside (with the exception of cyanidin), malvidin-3-(6-acetyl)-glucoside and
delphinidin, petunidin and malvidin-3-(6-p-coumaroyl)-glucosides, as well as the
guaiacyl-pyrano derivatives of malvidin-3-glucoside and its acylated forms have
been later confirmed in wines (Alcalde-Eon et al. 2004, 2006; Monagas et al. 2003;
Wang et al. 2003a; Pozo-Bay ́on et al. 2004; Boido et al. 2006).
Factors affecting the reaction.Schwarz, Hofmann, and Winterhalter (2004) have
investigated the factors influencing the formation of Pinotin A and its correlation
with wine age. It was found that the formation of this pigment was more depen-
dent on the concentration of caffeic acid than in that of malvidin-3-glucoside.
Although an exponential increase of the concentration of Pinotin A was observed
with prolonged aging time, the most rapid synthesis was observed when malvidin-
3-glucoside was degraded to a larger extent (2.5–4 year old wines) due to its partic-
ipation in other chemical reactions. This led to an increase of the ratio of caffeic