9B Flavanols, Flavonols and Dihydroflavonols 491
(Poncet-Legrand et al. 2006) and human salivary PRP (Pascal et al. 2007) occurs in
three stages:
- ligand binding and saturation of binding sites together with folding of the protein,
in the case of intrinsically unstructured proteins such as salivary PRP (Pascal
et al. 2007) or casein (Jobstl et al. 2006)- formation of rather small and homogenous protein-flavonoid aggregates
- further aggregation and precipitation
Adsorption of flavonols on PVPP also involves Van der Waals interactions with
associated hydrophobic effect and hydrogen bonding (Laborde et al. 2006).
Evidence in Wine
Protein haze due to interactions of flavanols with proteins and peptides is well
documented in beer (Outtrup 1989) but also takes place in wine. Pathogenesis
related proteins which are synthesized by vine following pathogen attacks seem
to be particularly involved in these processes (Waters et al. 1996). Protein precip-
itation also occurs as a result of fining treatments that consist in adding exoge-
nous proteins to precipitate tannins out inorder to stabilise the wine and reduce
its astringency. The proteins most commonly used for red wine fining are gelatins,
albumins and caseins. Plant proteins from lupine or wheat have recently been tested
as alternatives to gelatin (Maury et al. 2003). All of them, as well as salivary pro-
teins (Sarni-Manchado and Cheynier 2002) selectively precipitate higher molecular
weight flavanols ( Ricardo da Silva et al. 1991b; Maury et al. 2001; Sarni-Manchado
et al. 1999), which also exhibit higher astringency (Vidal et al. 2003). However, only
a small proportion of flavonoids was recovered in the pellet and the wine composi-
tion was not significantly modified by the treatment (Maury et al. 2003). The loss of
astringency observed after fining may thus be partly due to the inclusion of flavanols
in soluble complexes.
Factors Affecting the Interaction
Interactions and formation of insoluble complexes with proteins increase with
the number of phenolic rings, and especially ofo-diphenolic rings and thus with
polymerisation and galloylation (Haslam and Lilley 1988; McManus et al. 1985).
The intensity of mass spectrometry signals corresponding to soluble peptide fla-
vanol complexes increased from the monomers to the dimers and with galloylation
(Sarni-Manchado and Cheynier 2002). ITC experiments failed to detect any inter-
action of poly-L-proline with catechin or epicatechin while association constants
of 3. 7 × 104 /Mand8. 1 × 104 /M were determined for epigallocatechin gallate
and epicatechin gallate, respectively. Thatof an oligomeric procyanidin fraction
from grape seeds was even higher (3. 4 × 105 /M) confirming that the affinity of
flavanols for proteins increases with their chain length. Furthermore, interaction of